---------------------------------------------- # MEDLINE ID: 8371971 # MOTIF: [JP] MA0086.1 sna ---------------------------------------------- Nucleic Acids Res. 1993 Aug 25;21(17):3951-7. Definition of the DNA-binding site repertoire for the Drosophila transcription factor SNAIL. Mauhin V, Lutz Y, Dennefeld C, Alberga A. Laboratoire de Genetique Moleculaire des Eucaryotes du CNRS, Unite 184 de Biologie Moleculaire et de Genie Genetique de l'INSERM, Faculte de Medecine, Strasbourg, France. The Drosophila gene snail (sna) which encodes a zinc finger protein is essential for dorsal-ventral pattern formation in the developing embryo. We have defined a repertoire of SNAIL (SNA) binding sites using recombinant SNA proteins to select specific binding sequences from a pool of random sequence nucleotides. The bound sequences which were selected by multiple rounds of gel retardation and amplification by the polymerase chain reaction (PCR) were subsequently cloned and sequenced. The consensus sequence, 5'G/A A/t G/A A CAGGTG C/t A C 3', with a highly conserved core of 6 bases, CAGGTG, shares no significant homology with known binding sequences of other Drosophila zinc finger proteins. However, the CAGGTG core is identical to the core motif of aHLH (helix-loop-helix) binding sites. The strongest SNA binding is obtained with sequences containing this core motif whereas reduced binding is seen for sequences with canonical CANNTG HLH motifs. Interestingly, SNA binding is detected in the promoter region of the snail gene. Transient expression in co-transfection experiments using a SNA binding element (SBE) linked to a heterologous promoter indicates that SNA has the ability to function as a transcription activator. Publication Types: Research Support, Non-U.S. Gov't PMID: 8371971 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 2243767 # MOTIF: [JP] MA0083.1 SRF # MOTIF: [JP] MA0099.1 Fos ---------------------------------------------- Nucleic Acids Res. 1990 Nov 11;18(21):6197-204. A sensitive method for the determination of protein-DNA binding specificities. Pollock R, Treisman R. Imperial Cancer Research Fund Laboratories, Lincoln's Inn Fields, London, UK. We describe a sensitive and rapid method for determination of the sequence specificity of DNA binding proteins. The method allows recovery of specific sites using the small amounts of protein present in crude cell extracts or produced by cell-free translation reactions. Extract proteins are incubated with a pool of random sequence oligonucleotides, complexes purified by immunoprecipitation, and bound DNA amplified by the Polymerase Chain Reaction (PCR). This DNA is then used in further rounds of binding, immunoprecipitation, and amplification, until specific binding is detectable. With the transcription factor SRF as a model system, we demonstrate that authentic high affinity binding sites are recovered, and show that epitope tagging can be used to allow recovery of sites when specific antibodies are unavailable. We also show that specific sites bound by the Fos protein, which binds DNA with high affinity only when complexed with other polypeptides, are easily recovered by this technique. PMID: 2243767 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8543152 # MOTIF: [JP] MA0151.1 ARID3A ---------------------------------------------- Genes Dev. 1995 Dec 15;9(24):3067-82. The immunoglobulin heavy-chain matrix-associating regions are bound by Bright: a B cell-specific trans-activator that describes a new DNA-binding protein family. Herrscher RF, Kaplan MH, Lelsz DL, Das C, Scheuermann R, Tucker PW. Department of Microbiology, University of Texas Southwestern Medical Center, Dallas 75235-9048, USA. B lymphocyte-restricted transcription of immunoglobulin heavy-chain (IgH) genes is specified by elements within the variable region (VH) promoter and the intronic enhancer (E mu). The gene encoding a protein that binds a VH promoter proximal site necessary for induced mu-heavy-chain transcription has been cloned. This B-cell specific protein, termed Bright (B cell regulator of IgH transcription), is found in both soluble and matrix insoluble nuclear fractions. Bright binds the minor groove of a restricted ATC sequence that is sufficient for nuclear matrix association. This sequence motif is present in previously described matrix-associating regions (MARs) proximal to the promoter and flanking E mu. Bright can activate E mu-driven transcription by binding these sites, but only when they occur in their natural context and in cell lines permissive for E mu activity. To bind DNA, Bright requires a novel tetramerization domain and a previously undescribed domain that shares identity with several proteins, including SWI1, a component of the SWI/SNF complex. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8543152 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1748287 # MOTIF: [JP] MA0052.1 MEF2A ---------------------------------------------- Genes Dev. 1991 Dec;5(12A):2327-41. Human SRF-related proteins: DNA-binding properties and potential regulatory targets. Pollock R, Treisman R. Transcription Laboratory, Imperial Cancer Research Fund Laboratories, London, UK. Serum response factor (SRF) is a transcription factor that binds the sequence CC(A/T)6GG found in a number of growth factor-inducible and muscle-specific promoters. We describe the isolation and characterization of cDNA clones encoding a family of three human SRF-related DNA-binding proteins. Each of these RSRF (related to SRF) proteins contains an 86-amino-acid amino-terminal region related to the SRF DNA-binding domain: In RSRFC4 and RSRFC9, this region is identical, whereas that present in RSRFR2 differs by seven conservative amino acid substitutions. The DNA-binding specificity of the RSRF proteins, which recognize the consensus sequence CTA(A/T)4TAG, is distinct from that of SRF. The entire RSRF common region is required for DNA binding, and the differential sequence specificity of the RSRFs and SRF is the result of differences in the basic amino-terminal part of this domain. The RSRF proteins bind DNA as dimers and can dimerize with one another but not with SRF. Although the RSRF mRNAs are expressed in many cell types, RSRFR2 mRNA is expressed at elevated levels in several B-cell lines. Consistent with this, extracts from many cell types form CTA(A/T)4TAG-binding complexes that contain RSRF proteins, and oligonucleotides containing RSRF-binding sites function as promoter elements in transfection assays. Like SRF-binding sites, RSRF-binding sites are found in the regulatory sequences of a number of growth factor-inducible and muscle-specific genes, and we show that RSRF polypeptides are components of previously characterized binding activities that interact with these elements. We discuss the potential role of RSRF proteins in the regulation of these genes. PMID: 1748287 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9421508 # MOTIF: [JP] MA0089.1 NFE2L1::MafG ---------------------------------------------- Nucleic Acids Res. 1998 Jan 15;26(2):512-20. Interaction of the CNC-bZIP factor TCF11/LCR-F1/Nrf1 with MafG: binding-site selection and regulation of transcription. Johnsen O, Murphy P, Prydz H, Kolsto AB. Biotechnology Centre of Oslo, University of Oslo, PO Box 1125, Blindern N-0316 Oslo, Norway. We have previously shown that the widely expressed human transcription factor TCF11/LCR-F1/Nrf1 interacts with small Maf proteins and binds to a subclass of AP1-sites. Such sites are required for beta-globin 5' DNase I hypersensitive site 2 enhancer activity, erythroid porphobilinogen deaminase inducibility, hemin responsiveness by heme-oxygenase 1 and expression of the gene NAD(P)H:quinone oxidoreductase1. Here we report the optimal DNA-binding sequences for TCF11/LCR-F1/Nrf1 alone and as a heterodimer with MafG, identified by using binding-site selection. The heterodimer recognises a 5'-TGCTgaGTCAT-3' binding-site that is identical to the established NF-E2-site, the antioxidant response element and the heme-responsive element while the binding specificity of the homomer is less stringent. To investigate the activity of TCF11 through this selected site, both alone and in the presence of MafG, we have used a transient transfection assay. TCF11 alone activates transcription while MafG alone acts as a repressor. When co-expressed, MafG interferes with TCF11 transactivation in a dose dependent manner. This indicates that MafG protein, which heterodimerises efficiently with TCF11 in vitro (the heterodimer having a higher affinity for DNA than TCF11 alone), does not co-operate with TCF11 in transactivating transcription. We propose that since both these factors are widely expressed, they may act together to contribute to the negative regulation of this specific target site. Efficient positive regulation by TCF11 may require alternative partners with perhaps more restricted expression patterns. Publication Types: Research Support, Non-U.S. Gov't PMID: 9421508 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8253077 # MOTIF: [JP] MA0008.1 HAT5 ---------------------------------------------- EMBO J. 1993 Sep;12(9):3507-17. The Athb-1 and -2 HD-Zip domains homodimerize forming complexes of different DNA binding specificities. Sessa G, Morelli G, Ruberti I. Dipartimento di Genetica e Biologia Molecolare, Universita di Roma La Sapienza, Italy. The Arabidopsis Athb-1 and -2 proteins are characterized by the presence of a homeodomain (HD) with a closely linked leucine zipper motif (Zip). We have suggested that the HD-Zip motif could, via dimerization of the leucine zippers, recognize dyad-symmetric DNA sequences. Here we report an analysis of the DNA binding properties of the Athb-1 homeodomain-leucine zipper (HD-Zip-1) domain in vitro. DNA binding analysis performed using random-sequence DNA templates showed that the HD-Zip-1 domain, but not the Athb-1 HD alone, binds to DNA. The HD-Zip-1 domain recognizes a 9 bp dyad-symmetric sequence [CAAT(A/T)ATTG], as determined by selecting high-affinity binding sites from random-sequence DNA. Gel retardation assays demonstrated that the HD-Zip-1 domain binds to DNA as a dimer. Moreover, the analysis of the DNA binding activity of Athb-1 derivatives indicated that a correct spatial relationship between the HD and the Zip is essential for DNA binding. Finally, we determined that the Athb-2 HD-Zip domain recognizes a distinct 9 bp dyad-symmetric sequence [CAAT(G/C)ATTG]. A model of DNA binding by the HD-Zip proteins is proposed. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8253077 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15066426 # MOTIF: [JP] MF0001.1 ETS class # MOTIF: [JP] MF0002.1 bZIP CREB/G-box-like subclass # MOTIF: [JP] MF0003.1 REL class # MOTIF: [JP] MF0004.1 Nuclear Receptor class # MOTIF: [JP] MF0005.1 Forkhead class # MOTIF: [JP] MF0006.1 bZIP cEBP-like subclass # MOTIF: [JP] MF0007.1 bHLH(zip) class # MOTIF: [JP] MF0008.1 MADS class # MOTIF: [JP] MF0009.1 TRP(MYB) class # MOTIF: [JP] MF0010.1 Homeobox class # MOTIF: [JP] MF0011.1 HMG class ---------------------------------------------- J Mol Biol. 2004 Apr 23;338(2):207-15. Constrained binding site diversity within families of transcription factors enhances pattern discovery bioinformatics. Sandelin A, Wasserman WW. Center for Genomics and Bioinformatics, Karolinska Institutet, Stockholm, Sweden. Diverse computational and experimental efforts are required to elucidate the control circuitry regulating the transcription of human genes. The fusion of gene-specific promoter analyses with large microarray studies and bioinformatics advances has produced optimism that significant progress can be made in unravelling this complex network. Within bioinformatics, past emphasis for improved pattern discovery has been placed upon "phylogenetic footprinting", the identification of sequences conserved over moderate periods of evolution (e.g. human and mouse comparisons). We introduce a new direction in bioinformatics based on the constraints imposed by the structures of DNA-binding proteins. For most structurally related families of transcription factors, there are clear similarities in the sequences of the sites to which they bind. On the basis of this observation, we construct familial binding profiles for well-characterized transcription factor families. The profiles are shown to classify correctly the structural class of mediating transcription factors for novel motifs in 88% of cases. By incorporating the familial profiles into pattern discovery procedures, we demonstrate that functional binding sites can be found in genomic sequences of dramatically greater length than is possible otherwise. Thus, incorporating familial models can overcome the signal-to-noise challenge that has hindered the transition from microarray data to regulatory control sequences for human genes. Biochemically motivated constraints upon sequence diversity of binding sites will complement the genetically motivated constraints imposed in "phylogenetic footprinting" algorithms. PMID: 15066426 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19305498 # MOTIF: [JP] MA0149.1 EWSR1-FLI1 ---------------------------------------------- PLoS One. 2009;4(3):e4932. Epub 2009 Mar 23. The oncogenic EWS-FLI1 protein binds in vivo GGAA microsatellite sequences with potential transcriptional activation function. Guillon N, Tirode F, Boeva V, Zynovyev A, Barillot E, Delattre O. Institut Curie, Paris, France. The fusion between EWS and ETS family members is a key oncogenic event in Ewing tumors and important EWS-FLI1 target genes have been identified. However, until now, the search for EWS-FLI1 targets has been limited to promoter regions and no genome-wide comprehensive analysis of in vivo EWS-FLI1 binding sites has been undertaken. Using a ChIP-Seq approach to investigate EWS-FLI1-bound DNA sequences in two Ewing cell lines, we show that this chimeric transcription factor preferentially binds two types of sequences including consensus ETS motifs and microsatellite sequences. Most bound sites are found outside promoter regions. Microsatellites containing more than 9 GGAA repeats are very significantly enriched in EWS-FLI1 immunoprecipitates. Moreover, in reporter gene experiments, the transcription activation is highly dependent upon the number of repeats that are included in the construct. Importantly, in vivo EWS-FLI1-bound microsatellites are significantly associated with EWS-FLI1-driven gene activation. Put together, these results point out the likely contribution of microsatellite elements to long-distance transcription regulation and to oncogenesis. Publication Types: Research Support, Non-U.S. Gov't PMID: 19305498 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12101405 # MOTIF: [JP] MA0161.1 NFIC ---------------------------------------------- Nat Biotechnol. 2002 Aug;20(8):831-5. Epub 2002 Jul 8. High-throughput SELEX SAGE method for quantitative modeling of transcription-factor binding sites. Roulet E, Busso S, Camargo AA, Simpson AJ, Mermod N, Bucher P. Laboratory of Molecular Biotechnology, Center for Biotechnology UNIL-EPFL, and Institute of Animal Biology, University of Lausanne, 1015 Lausanne, Switzerland. The ability to determine the location and relative strength of all transcription-factor binding sites in a genome is important both for a comprehensive understanding of gene regulation and for effective promoter engineering in biotechnological applications. Here we present a bioinformatically driven experimental method to accurately define the DNA-binding sequence specificity of transcription factors. A generalized profile was used as a predictive quantitative model for binding sites, and its parameters were estimated from in vitro-selected ligands using standard hidden Markov model training algorithms. Computer simulations showed that several thousand low- to medium-affinity sequences are required to generate a profile of desired accuracy. To produce data on this scale, we applied high-throughput genomics methods to the biochemical problem addressed here. A method combining systematic evolution of ligands by exponential enrichment (SELEX) and serial analysis of gene expression (SAGE) protocols was coupled to an automated quality-controlled sequence extraction procedure based on Phred quality scores. This allowed the sequencing of a database of more than 10,000 potential DNA ligands for the CTF/NFI transcription factor. The resulting binding-site model defines the sequence specificity of this protein with a high degree of accuracy not achieved earlier and thereby makes it possible to identify previously unknown regulatory sequences in genomic DNA. A covariance analysis of the selected sites revealed non-independent base preferences at different nucleotide positions, providing insight into the binding mechanism. Publication Types: Research Support, Non-U.S. Gov't PMID: 12101405 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19111667 # MOTIF: [JP] MA0265.1 ABF1 # MOTIF: [JP] MA0266.1 ABF2 # MOTIF: [JP] MA0267.1 ACE2 # MOTIF: [JP] MA0268.1 ADR1 # MOTIF: [JP] MA0270.1 AFT2 # MOTIF: [JP] MA0275.1 ASG1 # MOTIF: [JP] MA0277.1 AZF1 # MOTIF: [JP] MA0280.1 CAT8 # MOTIF: [JP] MA0281.1 CBF1 # MOTIF: [JP] MA0282.1 CEP3 # MOTIF: [JP] MA0283.1 CHA4 # MOTIF: [JP] MA0284.1 CIN5 # MOTIF: [JP] MA0285.1 CRZ1 # MOTIF: [JP] MA0286.1 CST6 # MOTIF: [JP] MA0288.1 CUP9 # MOTIF: [JP] MA0289.1 DAL80 # MOTIF: [JP] MA0291.1 DAL82 # MOTIF: [JP] MA0292.1 ECM22 # MOTIF: [JP] MA0293.1 ECM23 # MOTIF: [JP] MA0294.1 EDS1 # MOTIF: [JP] MA0295.1 FHL1 # MOTIF: [JP] MA0297.1 FKH2 # MOTIF: [JP] MA0298.1 FZF1 # MOTIF: [JP] MA0300.1 GAT1 # MOTIF: [JP] MA0301.1 GAT3 # MOTIF: [JP] MA0302.1 GAT4 # MOTIF: [JP] MA0306.1 GIS1 # MOTIF: [JP] MA0307.1 GLN3 # MOTIF: [JP] MA0309.1 GZF3 # MOTIF: [JP] MA0310.1 HAC1 # MOTIF: [JP] MA0311.1 HAL9 # MOTIF: [JP] MA0312.1 HAP1 # MOTIF: [JP] MA0317.1 HCM1 # MOTIF: [JP] MA0318.1 HMRA2 # MOTIF: [JP] MA0319.1 HSF1 # MOTIF: [JP] MA0324.1 LEU3 # MOTIF: [JP] MA0325.1 LYS14 # MOTIF: [JP] MA0329.1 MBP1 # MOTIF: [JP] MA0333.1 MET31 # MOTIF: [JP] MA0334.1 MET32 # MOTIF: [JP] MA0337.1 MIG1 # MOTIF: [JP] MA0338.1 MIG2 # MOTIF: [JP] MA0339.1 MIG3 # MOTIF: [JP] MA0341.1 MSN2 # MOTIF: [JP] MA0342.1 MSN4 # MOTIF: [JP] MA0344.1 NHP10 # MOTIF: [JP] MA0348.1 OAF1 # MOTIF: [JP] MA0352.1 PDR1 # MOTIF: [JP] MA0354.1 PDR8 # MOTIF: [JP] MA0355.1 PHD1 # MOTIF: [JP] MA0356.1 PHO2 # MOTIF: [JP] MA0358.1 PUT3 # MOTIF: [JP] MA0359.1 RAP1 # MOTIF: [JP] MA0360.1 RDR1 # MOTIF: [JP] MA0361.1 RDS1 # MOTIF: [JP] MA0362.1 RDS2 # MOTIF: [JP] MA0363.1 REB1 # MOTIF: [JP] MA0364.1 REI1 # MOTIF: [JP] MA0365.1 RFX1 # MOTIF: [JP] MA0366.1 RGM1 # MOTIF: [JP] MA0367.1 RGT1 # MOTIF: [JP] MA0368.1 RIM101 # MOTIF: [JP] MA0371.1 ROX1 # MOTIF: [JP] MA0372.1 RPH1 # MOTIF: [JP] MA0373.1 RPN4 # MOTIF: [JP] MA0374.1 RSC3 # MOTIF: [JP] MA0375.1 RSC30 # MOTIF: [JP] MA0379.1 SIG1 # MOTIF: [JP] MA0380.1 SIP4 # MOTIF: [JP] MA0381.1 SKN7 # MOTIF: [JP] MA0385.1 SOK2 # MOTIF: [JP] MA0389.1 SRD1 # MOTIF: [JP] MA0391.1 STB4 # MOTIF: [JP] MA0392.1 STB5 # MOTIF: [JP] MA0396.1 STP3 # MOTIF: [JP] MA0397.1 STP4 # MOTIF: [JP] MA0398.1 SUM1 # MOTIF: [JP] MA0401.1 SWI4 # MOTIF: [JP] MA0402.1 SWI5 # MOTIF: [JP] MA0403.1 TBF1 # MOTIF: [JP] MA0404.1 TBS1 # MOTIF: [JP] MA0405.1 TEA1 # MOTIF: [JP] MA0406.1 TEC1 # MOTIF: [JP] MA0408.1 TOS8 # MOTIF: [JP] MA0409.1 TYE7 # MOTIF: [JP] MA0410.1 UGA3 # MOTIF: [JP] MA0411.1 UPC2 # MOTIF: [JP] MA0414.1 XBP1 # MOTIF: [JP] MA0416.1 YAP3 # MOTIF: [JP] MA0420.1 YBR239C # MOTIF: [JP] MA0422.1 YDR520C # MOTIF: [JP] MA0423.1 YER130C # MOTIF: [JP] MA0424.1 YER184C # MOTIF: [JP] MA0425.1 YGR067C # MOTIF: [JP] MA0427.1 YJL103C # MOTIF: [JP] MA0428.1 YKL222C # MOTIF: [JP] MA0429.1 YLL054C # MOTIF: [JP] MA0430.1 YLR278C # MOTIF: [JP] MA0431.1 YML081W # MOTIF: [JP] MA0432.1 YNR063W # MOTIF: [JP] MA0433.1 YOX1 # MOTIF: [JP] MA0434.1 YPR013C # MOTIF: [JP] MA0436.1 YPR022C # MOTIF: [JP] MA0437.1 YPR196W # MOTIF: [JP] MA0438.1 YRM1 # MOTIF: [JP] MA0439.1 YRR1 # MOTIF: [JP] MA0441.1 ZMS1 ---------------------------------------------- Mol Cell. 2008 Dec 26;32(6):878-87. A library of yeast transcription factor motifs reveals a widespread function for Rsc3 in targeting nucleosome exclusion at promoters. Badis G, Chan ET, van Bakel H, Pena-Castillo L, Tillo D, Tsui K, Carlson CD, Gossett AJ, Hasinoff MJ, Warren CL, Gebbia M, Talukder S, Yang A, Mnaimneh S, Terterov D, Coburn D, Li Yeo A, Yeo ZX, Clarke ND, Lieb JD, Ansari AZ, Nislow C, Hughes TR. Banting and Best Department of Medical Research, University of Toronto, Toronto, ON M5S 3E1, Canada. The sequence specificity of DNA-binding proteins is the primary mechanism by which the cell recognizes genomic features. Here, we describe systematic determination of yeast transcription factor DNA-binding specificities. We obtained binding specificities for 112 DNA-binding proteins representing 19 distinct structural classes. One-third of the binding specificities have not been previously reported. Several binding sequences have striking genomic distributions relative to transcription start sites, supporting their biological relevance and suggesting a role in promoter architecture. Among these are Rsc3 binding sequences, containing the core CGCG, which are found preferentially approximately 100 bp upstream of transcription start sites. Mutation of RSC3 results in a dramatic increase in nucleosome occupancy in hundreds of proximal promoters containing a Rsc3 binding element, but has little impact on promoters lacking Rsc3 binding sequences, indicating that Rsc3 plays a broad role in targeting nucleosome exclusion at yeast promoters. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 19111667 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8636240 # MOTIF: [JP] MA0078.1 Sox17 ---------------------------------------------- J Cell Biol. 1996 May;133(3):667-81. Identification of two Sox17 messenger RNA isoforms, with and without the high mobility group box region, and their differential expression in mouse spermatogenesis. Kanai Y, Kanai-Azuma M, Noce T, Saido TC, Shiroishi T, Hayashi Y, Yazaki K. Department of Ultrastructural Research, Tokyo Metropolitan Institute of Medical Science, Japan. The different mRNA isoforms of the mouse Sox17 gene were isolated from adult mouse testis cDNAs. One form (referred to as form Sox17) encodes an Sry-related protein of 419 amino acids containing a single high mobility group box near the NH2-terminus, while the other form (referred to as form t-Sox17) shows a unique mRNA isoform of the Sox17 gene with a partial deletion of the HMG box region. Analysis of genomic DNA revealed that these two isoforms were produced at least by alternative splicing of the exon corresponding to the 5' untranslated region and NH2-terminal 102 amino acids. RNA analyses in the testis revealed that form Sox17 began at the pachytene spermatocyte stage and was highly accumulated in round spermatids. Protein analyses revealed that t-Sox17 isoforms, as well as Sox17 isoforms, were translated into the protein products in the testis, although the amount of t-Sox17 products is lower in comparison to the high accumulation of t-Sox17 mRNA. By the electrophoretic mobility-shift assay and the random selection assay using recombinant Sox17 and t-Sox17 proteins, Sox17 protein is a DNA-binding protein with a similar sequence specificity to Sry and the other members of Sox family proteins, while t-Sox17 shows no apparent DNA-binding activity. Moreover, by a cotransfection experiment using a luciferase reporter gene, Sox17 could stimulate transcription through its binding site, but t-Sox17 had little effect on reporter gene expression. Thus, these findings suggest that Sox17 may function as a transcriptional activator in the premeiotic germ cells, and that a splicing switch into t-Sox17 may lead to the loss of its function in the postmeiotic germ cells. Publication Types: Research Support, Non-U.S. Gov't PMID: 8636240 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18981474 # MOTIF: [JP] MA0065.2 PPARG::RXRA ---------------------------------------------- Genes Dev. 2008 Nov 1;22(21):2953-67. Genome-wide profiling of PPARgamma:RXR and RNA polymerase II occupancy reveals temporal activation of distinct metabolic pathways and changes in RXR dimer composition during adipogenesis. Nielsen R, Pedersen TA, Hagenbeek D, Moulos P, Siersbaek R, Megens E, Denissov S, Borgesen M, Francoijs KJ, Mandrup S, Stunnenberg HG. Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense M, Denmark. The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) is a key regulator of adipocyte differentiation in vivo and ex vivo and has been shown to control the expression of several adipocyte-specific genes. In this study, we used chromatin immunoprecipitation combined with deep sequencing to generate genome-wide maps of PPARgamma and retinoid X receptor (RXR)-binding sites, and RNA polymerase II (RNAPII) occupancy at very high resolution throughout adipocyte differentiation of 3T3-L1 cells. We identify >5000 high-confidence shared PPARgamma:RXR-binding sites in adipocytes and show that during early stages of differentiation, many of these are preoccupied by non-PPARgamma RXR-heterodimers. Different temporal and compositional patterns of occupancy are observed. In addition, we detect co-occupancy with members of the C/EBP family. Analysis of RNAPII occupancy uncovers distinct clusters of similarly regulated genes of different biological processes. PPARgamma:RXR binding is associated with the majority of induced genes, and sites are particularly abundant in the vicinity of genes involved in lipid and glucose metabolism. Our analyses represent the first genome-wide map of PPARgamma:RXR target sites and changes in RNAPII occupancy throughout adipocyte differentiation and indicate that a hitherto unrecognized high number of adipocyte genes of distinctly regulated pathways are directly activated by PPARgamma:RXR. Publication Types: Research Support, Non-U.S. Gov't PMID: 18981474 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 2192357 # MOTIF: [JP] MA0079.1 SP1 ---------------------------------------------- Nucleic Acids Res. 1990 Jun 11;18(11):3203-9. Target Detection Assay (TDA): a versatile procedure to determine DNA binding sites as demonstrated on SP1 protein. Thiesen HJ, Bach C. Basel Institute for Immunology, Switzerland. We developed a rapid method designated Target Detection Assay (TDA) to determine DNA binding sites for putative DNA binding proteins. A purified, functionally active DNA binding protein and a pool of random double-stranded oligonucleotides harbouring PCR primer sites at each end are included the TDA cycle which consists of four separate steps: a DNA protein incubation step, a protein DNA complex separation step, a DNA elution step and a polymerase chain reaction (PCR) DNA amplification step. The stringency of selection can be increased in consecutive TDA cycles. Since tiny amounts of retained DNA can be rescued by PCR, buffer systems, salt concentrations and competitor DNA contents can be varied in order to determine high affinity binding sites for the protein of choice. To test the efficiency of the TDA procedure potential DNA binding sites were selected by the DNA binding protein SP1 from a pool of oligonucleotides with random nucleotides at 12 positions. Target sites selected by recombinant SP1 closely matched the SP1 consensus site. If DNA recognition sites have to be determined for known, mutated or putative DNA binding proteins, the Target Detection Assay (TDA) is a versatile and rapid technique for consideration. Publication Types: Research Support, Non-U.S. Gov't PMID: 2192357 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9009278 # MOTIF: [JP] MA0088.1 znf143 ---------------------------------------------- EMBO J. 1997 Jan 2;16(1):173-81. Staf, a promiscuous activator for enhanced transcription by RNA polymerases II and III. Schaub M, Myslinski E, Schuster C, Krol A, Carbon P. UPR 9002 du CNRS Structure des Macromolecules Biologiques et Mecanismes de Reconnaissance, IBMC, Strasbourg, France. Staf is a zinc finger protein that we recently identified as the transcriptional activator of the RNA polymerase III-transcribed selenocysteine tRNA gene. In this work we demonstrate that enhanced transcription of the majority of vertebrate snRNA and snRNA-type genes, transcribed by RNA polymerases II and III, also requires Staf. DNA binding assays and microinjection of mutant genes into Xenopus oocytes showed the presence of Staf-responsive elements in the genes for human U4C, U6, Y4 and 7SK, Xenopus U1b1, U2, U5 and MRP and mouse U6 RNAs. Using recombinant Staf, we established that it mediates the activating properties of Staf-responsive elements on RNA polymerase II and III snRNA promoters in vivo. Lastly a 19 bp consensus sequence for the Staf binding site, YY(A/T)CCC(A/G)N(A/C)AT(G/C)C(A/C)YY-RCR, was derived by binding site selection. It enabled us to identify 23 other snRNA and snRNA-type genes carrying potential Staf binding sites. Altogether, our results emphasize the prime importance of Staf as a novel activator for enhanced transcription of snRNA and snRNA-type genes. Publication Types: Research Support, Non-U.S. Gov't PMID: 9009278 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1411535 # MOTIF: [JP] MA0024.1 E2F1 ---------------------------------------------- Science. 1992 Oct 16;258(5081):424-9. E2F: a link between the Rb tumor suppressor protein and viral oncoproteins. Nevins JR. Howard Hughes Medical Institute, Section of Genetics, Duke University Medical Center, Durham, NC 27710. The cellular transcription factor E2F, previously identified as a component of early adenovirus transcription, has now been shown to be important in cell proliferation control. E2F appears to be a functional target for the action of the tumor suppressor protein Rb that is encoded by the retinoblastoma susceptibility gene. The disruption of this E2F-Rb interaction, as well as a complex involving E2F in association with the cell cycle-regulated cyclin A-cdk2 kinase complex, may be a common mechanism of action for the oncoproteins encoded by the DNA tumor viruses. PMID: 1411535 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16314527 # MOTIF: [JP] MA0261.1 lin-14 ---------------------------------------------- Mol Cell Biol. 2005 Dec;25(24):11059-72. The Caenorhabditis elegans heterochronic regulator LIN-14 is a novel transcription factor that controls the developmental timing of transcription from the insulin/insulin-like growth factor gene ins-33 by direct DNA binding. Hristova M, Birse D, Hong Y, Ambros V. Dartmouth Medical School, Department of Genetics, Hanover, NH 03755, USA. A temporal gradient of the novel nuclear protein LIN-14 specifies the timing and sequence of stage-specific developmental events in Caenorhabditis elegans. The profound effects of lin-14 mutations on worm development suggest that LIN-14 directly or indirectly regulates stage-specific gene expression. We show that LIN-14 can associate with chromatin in vivo and has in vitro DNA binding activity. A bacterially expressed C-terminal domain of LIN-14 was used to select DNA sequences that contain a putative consensus binding site from a pool of randomized double-stranded oligonucleotides. To identify candidates for genes directly regulated by lin-14, we employed DNA microarray hybridization to compare the mRNA abundance of C. elegans genes in wild-type animals to that in mutants with reduced or elevated lin-14 activity. Five of the candidate LIN-14 target genes identified by microarrays, including the insulin/insulin-like growth factor family gene ins-33, contain putative LIN-14 consensus sites in their upstream DNA sequences. Genetic analysis indicates that the developmental regulation of ins-33 mRNA involves the stage-specific repression of ins-33 transcription by LIN-14 via sequence-specific DNA binding. These results reinforce the conclusion that lin-14 encodes a novel class of transcription factor. Publication Types: Research Support, N.I.H., Extramural PMID: 16314527 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17442748 # MOTIF: [JP] CN0001.1 LM1 # MOTIF: [JP] CN0002.1 LM2 # MOTIF: [JP] CN0003.1 LM3 # MOTIF: [JP] CN0004.1 LM4 # MOTIF: [JP] CN0005.1 LM5 # MOTIF: [JP] CN0006.1 LM6 # MOTIF: [JP] CN0007.1 LM7 # MOTIF: [JP] CN0008.1 LM8 # MOTIF: [JP] CN0009.1 LM9 # MOTIF: [JP] CN0010.1 LM10 # MOTIF: [JP] CN0011.1 LM11 # MOTIF: [JP] CN0012.1 LM12 # MOTIF: [JP] CN0013.1 LM13 # MOTIF: [JP] CN0014.1 LM14 # MOTIF: [JP] CN0015.1 LM15 # MOTIF: [JP] CN0016.1 LM16 # MOTIF: [JP] CN0017.1 LM17 # MOTIF: [JP] CN0018.1 LM18 # MOTIF: [JP] CN0019.1 LM19 # MOTIF: [JP] CN0020.1 LM20 # MOTIF: [JP] CN0021.1 LM21 # MOTIF: [JP] CN0022.1 LM22 # MOTIF: [JP] CN0023.1 LM23 # MOTIF: [JP] CN0024.1 LM24 # MOTIF: [JP] CN0025.1 LM25 # MOTIF: [JP] CN0026.1 LM26 # MOTIF: [JP] CN0027.1 LM27 # MOTIF: [JP] CN0028.1 LM28 # MOTIF: [JP] CN0029.1 LM29 # MOTIF: [JP] CN0030.1 LM30 # 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MOTIF: [JP] CN0208.1 LM208 # MOTIF: [JP] CN0209.1 LM209 # MOTIF: [JP] CN0210.1 LM210 # MOTIF: [JP] CN0211.1 LM211 # MOTIF: [JP] CN0212.1 LM212 # MOTIF: [JP] CN0213.1 LM213 # MOTIF: [JP] CN0214.1 LM214 # MOTIF: [JP] CN0215.1 LM215 # MOTIF: [JP] CN0216.1 LM216 # MOTIF: [JP] CN0217.1 LM217 # MOTIF: [JP] CN0218.1 LM218 # MOTIF: [JP] CN0219.1 LM219 # MOTIF: [JP] CN0220.1 LM220 # MOTIF: [JP] CN0221.1 LM221 # MOTIF: [JP] CN0222.1 LM222 # MOTIF: [JP] CN0223.1 LM223 # MOTIF: [JP] CN0224.1 LM224 # MOTIF: [JP] CN0225.1 LM225 # MOTIF: [JP] CN0226.1 LM226 # MOTIF: [JP] CN0227.1 LM227 # MOTIF: [JP] CN0228.1 LM228 # MOTIF: [JP] CN0229.1 LM229 # MOTIF: [JP] CN0230.1 LM230 # MOTIF: [JP] CN0231.1 LM231 # MOTIF: [JP] CN0232.1 LM232 # MOTIF: [JP] CN0233.1 LM233 ---------------------------------------------- Proc Natl Acad Sci U S A. 2007 Apr 24;104(17):7145-50. Epub 2007 Apr 18. Systematic discovery of regulatory motifs in conserved regions of the human genome, including thousands of CTCF insulator sites. Xie X, Mikkelsen TS, Gnirke A, Lindblad-Toh K, Kellis M, Lander ES. Broad Institute of MIT and Harvard, Massachusetts Institute of Technology and Harvard Medical School, Cambridge, MA 02142, USA. Conserved noncoding elements (CNEs) constitute the majority of sequences under purifying selection in the human genome, yet their function remains largely unknown. Experimental evidence suggests that many of these elements play regulatory roles, but little is known about regulatory motifs contained within them. Here we describe a systematic approach to discover and characterize regulatory motifs within mammalian CNEs by searching for long motifs (12-22 nt) with significant enrichment in CNEs and studying their biochemical and genomic properties. Our analysis identifies 233 long motifs (LMs), matching a total of approximately 60,000 conserved instances across the human genome. These motifs include 16 previously known regulatory elements, such as the histone 3'-UTR motif and the neuron-restrictive silencer element, as well as striking examples of novel functional elements. The most highly enriched motif (LM1) corresponds to the X-box motif known from yeast and nematode. We show that it is bound by the RFX1 protein and identify thousands of conserved motif instances, suggesting a broad role for the RFX family in gene regulation. A second group of motifs (LM2*) does not match any previously known motif. We demonstrate by biochemical and computational methods that it defines a binding site for the CTCF protein, which is involved in insulator function to limit the spread of gene activation. We identify nearly 15,000 conserved sites that likely serve as insulators, and we show that nearby genes separated by predicted CTCF sites show markedly reduced correlation in gene expression. These sites may thus partition the human genome into domains of expression. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 17442748 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15661650 # MOTIF: [JP] MA0118.1 Macho-1 ---------------------------------------------- Gene Expr Patterns. 2005 Feb;5(3):429-37. Macho-1 functions as transcriptional activator for muscle formation in embryos of the ascidian Halocynthia roretzi. Sawada K, Fukushima Y, Nishida H. Department of Biology, Graduate School of Science, Osaka University, 1-1 Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan. Various kinds of maternal factor that play crucial roles in embryogenesis are present and localized in the ascidian egg cytoplasm. Localized maternal mRNA of the macho-1 gene is a muscle determinant in the embryo of the ascidian Halocynthia roretzi. The macho-1 protein has a zinc-finger domain and accumulates in nuclei, being expected to function as a DNA-binding transcription factor. In the present study, we show that macho-1 is, indeed, a DNA-binding transcriptional activator, and directly or indirectly regulates the expression of six downstream genes. Macho-1 was required and sufficient for expression of the muscle actin, myosin, calcium transporter, myogenic factor, Tbx6, and snail genes, whose expression is initiated in muscle blastomeres at the cleavage stages in normal embryos. Furthermore, when macho-1 conjugated with a transcription-repression domain of Drosophila engrailed (En(R)) was expressed in embryos, it repressed expression of these downstream genes. In contrast, expression of macho-1 fused with a transcription-activation domain of VP16 caused ectopic expression of these muscle genes in non-muscle blastomeres. PCR-assisted binding-site selection and gel-retardation assay showed that macho-1 protein binds to the consensus target sequence (TGGGTGGTC) for GLI/Zic-family proteins, and that three guanine residues with underlines are crucial for the specificity. The 5' promoter region of the muscle actin gene supported expression of the reporter gene only in muscle cells at late stage. By contrast, when the target sequence was added to the promoter, it well reproduced early expression of muscle actin at the cleavage stage, indicating that macho-1 can recognize the target sequence in vivo. In conclusion, the maternal muscle determinant macho-1 functions as a transcription factor that positively regulates gene expression for muscle formation in ascidian embryos. Publication Types: Research Support, Non-U.S. Gov't PMID: 15661650 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10487868 # MOTIF: [JP] MA0304.1 GCR1 # MOTIF: [JP] MA0328.1 MATALPHA2 # MOTIF: [JP] MA0369.1 RLM1 ---------------------------------------------- Bioinformatics. 1999 Jul-Aug;15(7-8):607-11. SCPD: a promoter database of the yeast Saccharomyces cerevisiae. Zhu J, Zhang MQ. Cold Spring Harbor Laboratory, PO Box 100, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA. zhuj@cshl.org MOTIVATION: In order to facilitate a systematic study of the promoters and transcriptionally regulatory cis-elements of the yeast Saccharomyces cerevisiae on a genomic scale, we have developed a comprehensive yeast-specific promoter database, SCPD. RESULTS: Currently SCPD contains 580 experimentally mapped transcription factor (TF) binding sites and 425 transcriptional start sites (TSS) as its primary data entries. It also contains relevant binding affinity and expression data where available. In addition to mechanisms for promoter information (including sequence) retrieval and a data submission form, SCPD also provides some simple but useful tools for promoter sequence analysis. AVAILABILITY: SCPD can be accessed from the URL http://cgsigma.cshl.org/jian. The database is continually updated. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 10487868 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9927589 # MOTIF: [JP] MA0262.1 mab-3 ---------------------------------------------- Development. 1999 Feb;126(5):873-81. Similarity of DNA binding and transcriptional regulation by Caenorhabditis elegans MAB-3 and Drosophila melanogaster DSX suggests conservation of sex determining mechanisms. Yi W, Zarkower D. Biochemistry, Molecular Biology and Biophysics Graduate Program, and Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455, USA. Although most animals occur in two sexes, the molecular pathways they employ to control sexual development vary considerably. The only known molecular similarity between phyla in sex determination is between two genes, mab-3 from C. elegans, and doublesex (dsx) from Drosophila. Both genes contain a DNA binding motif called a DM domain and they regulate similar aspects of sexual development, including yolk protein synthesis and peripheral nervous system differentiation. Here we show that MAB-3, like the DSX proteins, is a direct regulator of yolk protein gene transcription. We show that despite containing different numbers of DM domains MAB-3 and DSX bind to similar DNA sequences. mab-3 mutations deregulate vitellogenin synthesis at the level of transcription, resulting in expression in both sexes, and the vitellogenin genes have potential MAB-3 binding sites upstream of their transcriptional start sites. MAB-3 binds to a site in the vit-2 promoter in vitro, and this site is required in vivo to prevent transcription of a vit-2 reporter construct in males, suggesting that MAB-3 is a direct repressor of vitellogenin transcription. This is the first direct link between the sex determination regulatory pathway and sex-specific structural genes in C. elegans, and it suggests that nematodes and insects use at least some of the same mechanisms to control sexual development. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 9927589 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10327073 # MOTIF: [JP] MA0119.1 TLX1::NFIC ---------------------------------------------- Oncogene. 1999 Apr 1;18(13):2273-9. HOX11 interacts with CTF1 and mediates hematopoietic precursor cell immortalization. Zhang N, Shen W, Hawley RG, Lu M. Department of Medicine, University of California, San Diego, La Jolla 92093-0671, USA. HOX11 is a homeodomain-containing oncogenic transcription factor that immortalizes hematopoietic precursor cells. The mechanism by which HOX11 facilitates this initial step of leukemogenesis is, however, not well understood. We have used a DNA binding site selection assay to investigate cooperative DNA binding by HOX11 with other transcription factors. A consensus sequence was derived and identified as the binding site for the CCAAT-box-binding transcription factors (CTF). HOX11 was shown to interact in vitro and in vivo with CTF1. Retrovirus-mediated transduction of an antisense CTF1 cDNA dramatically reduced the proliferative capacity of HOX11-immortalized hematopoietic precursor cells. CTF1 is, therefore, the first HOX11 protein partner identified that plays an important role in hematopoietic precursor cell immortalization. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 10327073 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 2507923 # MOTIF: [JP] MA0049.1 hb ---------------------------------------------- Nature. 1989 Sep 28;341(6240):331-5. Sequence-specific DNA-binding activities of the gap proteins encoded by hunchback and Kruppel in Drosophila. Stanojevic D, Hoey T, Levine M. Department of Biological Sciences, Fairchild Center, Columbia University, New York, New York 10027. The segmentation of the Drosophila body plan depends on a hierarchy of interactions among approximately 20-25 regulatory genes that are active in the early embryo (refs 1-4; for a review see ref. 5). The gap genes have a key role in this process and are responsible for the periodic expression of certain pair-rule genes and the localized expression of several homoeotic genes. The two best characterized gap genes, hunchback (hb) and Kruppel (Kr), contain homologies with the zinc-finger DNA-binding motif, although their mode of action in the early embryo is unknown. Here we report that both of the proteins encoded by these genes possess sequence-specific DNA-binding activities, which indicates that they might regulate gene expression at the level of transcription. The binding sites of the hb gene product are related by a 10-base pair (bp) consensus sequence, G/A C/C ATAAAAAA, whereas the binding sites of the Kr gene product share a distinct 10-bp motif, AACGGGTTAA. It is possible that the hb and Kr proteins cooperatively regulate gene expression, because they are expressed in broad, overlapping gradients in the early embryo. We also provide evidence that the on/off periodicity of the pair-rule gene even-skipped (eve) involves the interaction of the hb and Kr proteins with defined eve promoter elements. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 2507923 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9153225 # MOTIF: [JP] MA0042.1 FOXI1 ---------------------------------------------- J Biol Chem. 1997 May 23;272(21):13725-30. The winged helix transcriptional activator HFH-3 is expressed in the distal tubules of embryonic and adult mouse kidney. Overdier DG, Ye H, Peterson RS, Clevidence DE, Costa RH. Department of Biochemistry, University of Illinois, Chicago, Illinois 60612-7334, USA. The hepatocyte nuclear factor-3 (HNF-3)/fork head homolog (HFH) proteins are an extensive family of transcription factors, which share homology in the winged helix DNA binding domain. Members of the HFH/winged helix family have been implicated in cell fate determination during pattern formation, in organogenesis, and in cell-type-specific gene expression. In this study we isolated a full-length HFH-3 cDNA clone from a human kidney library which encoded a 351-amino acid protein containing a centrally located winged helix DNA binding domain. We demonstrate that HFH-3 is a potent transcriptional activator requiring 138 C-terminal residues for activity. We used in situ hybridization to demonstrate that HFH-3 expression is restricted to the epithelium of the renal distal convoluted tubules. We determined the HFH-3 DNA binding consensus sequence by in vitro DNA binding site selection using recombinant HFH-3 protein and used this consensus sequence to identify putative HFH-3 target genes expressed there. These putative HFH-3 target genes include the Na/K-ATPase, Na/H and anion exchangers, E-cadherin, and mineralocorticoid receptor genes as well as genes for the transcription factors HNF-1, vHNF-1, and HNF-4. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 9153225 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8383622 # MOTIF: [JP] MA0062.1 GABPA ---------------------------------------------- Genes Dev. 1993 Mar;7(3):380-92. Identity of GABP with NRF-2, a multisubunit activator of cytochrome oxidase expression, reveals a cellular role for an ETS domain activator of viral promoters. Virbasius JV, Virbasius CA, Scarpulla RC. Department of Cell, Molecular, and Structural Biology, Northwestern University Medical School, Chicago, Illinois 60611. The ETS domain proteins are a diverse family of transcriptional activators that have been implicated recently in the expression of a number of cell-specific and viral promoters. Nuclear respiratory factor 2 (NRF-2) is a nuclear transcription factor that activates the proximal promoter of the rat cytochrome c oxidase subunit IV (RCO4) gene through tandem sequence elements. These elements conform to the consensus for high-affinity ETS domain recognition sites. We have now purified NRF-2 to homogeneity from HeLa cells and find that it consists of five polypeptides, only one of which has intrinsic DNA-binding ability. The others participate in the formation of heteromeric complexes with distinct binding properties. NRF-2 also specifically recognizes multiple binding sites in the mouse cytochrome c oxidase subunit Vb (MCO5b) gene. As in the functionally related RCO4 gene, tandemly arranged NRF-2 sites are essential for the activity of the proximal MCO5b promoter, further substantiating a role for NRF-2 in respiratory chain expression. Determination of peptide sequences from the various subunits of HeLa NRF-2 reveals a high degree of sequence identity with mouse GA-binding protein (GABP), a multisubunit ETS domain activator of herpes simplex virus immediate early genes. A cellular role in the activation of nuclear genes specifying mitochondrial respiratory function is thus assigned to an ETS domain activator of viral promoters. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8383622 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19553195 # MOTIF: [JP] MA0047.2 Foxa2 ---------------------------------------------- Nucleic Acids Res. 2009 Sep;37(17):e113. Epub 2009 Jun 24. Extracting transcription factor targets from ChIP-Seq data. Tuteja G, White P, Schug J, Kaestner KH. Department of Genetics and Institute of Diabetes, Obesity and Metabolism, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA. ChIP-Seq technology, which combines chromatin immunoprecipitation (ChIP) with massively parallel sequencing, is rapidly replacing ChIP-on-chip for the genome-wide identification of transcription factor binding events. Identifying bound regions from the large number of sequence tags produced by ChIP-Seq is a challenging task. Here, we present GLITR (GLobal Identifier of Target Regions), which accurately identifies enriched regions in target data by calculating a fold-change based on random samples of control (input chromatin) data. GLITR uses a classification method to identify regions in ChIP data that have a peak height and fold-change which do not resemble regions in an input sample. We compare GLITR to several recent methods and show that GLITR has improved sensitivity for identifying bound regions closely matching the consensus sequence of a given transcription factor, and can detect bona fide transcription factor targets missed by other programs. We also use GLITR to address the issue of sequencing depth, and show that sequencing biological replicates identifies far more binding regions than re-sequencing the same sample. PMID: 19553195 [PubMed - in process] ---------------------------------------------- # MEDLINE ID: 7624145 # MOTIF: [JP] MA0080.1 SPI1 # MOTIF: [JP] MA0081.1 SPIB ---------------------------------------------- Oncogene. 1995 Jul 20;11(2):303-13. DNA binding specificities of Spi-1/PU.1 and Spi-B transcription factors and identification of a Spi-1/Spi-B binding site in the c-fes/c-fps promoter. Ray-Gallet D, Mao C, Tavitian A, Moreau-Gachelin F. Unite 248 INSERM, Faculte de Medecine Lariboisiere Saint-Louis, Paris, France. Spi-1/PU.1 and Spi-B encode hematopoietic-specific transcription factors that are the most distantly related members of the Ets family. The Ets proteins share a conserved 85 amino acids DNA binding domain, the Ets domain and recognize various DNA target sites around a common core 5'-GGAA/T-3'. The DNA binding specificities of Spi-1 and Spi-B were investigated by using the method of polymerase chain reaction (PCR)-mediated random site selection. The deduced Spi-1 and Spi-B consensus binding sites are very similar suggesting that the functional activities of Spi-1 and Spi-B cannot be distinguished on the basis of their DNA binding specificities. We identified a putative Spi-1/Spi-B binding site in the promoter region of the c-fes/c-fps protooncogene which encodes a tyrosine kinase expressed predominantly in myeloid cells. In vitro translated Spi-1 and Spi-B proteins were capable to bind this site similarly and to activate the c-fes promoter in HeLa transfected cells. We showed that Spi-1 binds the Spi-1/Spi-B binding site of c-fes in HL-60 cells suggesting that Spi-1 may be involved in the regulation of c-fes transcription in myeloid cells. Intriguingly, we detected only Spi-1 binding to this site in the Raji cell line which express both Spi-1 and Spi-B proteins. This suggests that Spi-1 and Spi-B exhibit different DNA binding activities in vivo although they share similar DNA binding specificities in vitro. Publication Types: Research Support, Non-U.S. Gov't PMID: 7624145 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10637336 # MOTIF: [JP] MA0126.1 ovo ---------------------------------------------- Nucleic Acids Res. 2000 Feb 1;28(3):826-34. Characterization of Drosophila OVO protein DNA binding specificity using random DNA oligomer selection suggests zinc finger degeneration. Lee S, Garfinkel MD. Division of Biology, Illinois Institute of Technology, Chicago, IL 60616, USA. The Drosophila melanogaster ovo locus codes for several tissue- and stage-specific proteins that all possess a common C-terminal array of four C(2)H(2)zinc fingers. Three fingers conform to the motif framework and are evolutionarily conserved; the fourth diverges considerably. The ovo genetic function affects germ cell viability, sex identity and oogenesis, while the overlapping svb function is a key selector for epidermal structures under the control of wnt and EGF receptor signaling. We isolated synthetic DNA oligomers bound by the OVO zinc finger array from a high complexity starting population and derived a statistically significant 9 bp long DNA consensus sequence, which is nearly identical to a consensus derived from several Drosophila genes known or suspected of being regulated by the ovo function in vivo. The DNA consensus recognized by Drosophila OVO protein is atypical for zinc finger proteins in that it does not conform to many of the 'rules' for the interaction of amino acid contact residues and DNA bases. Additionally, our results suggest that only three of the OVO zinc fingers contribute to DNA-binding specificity. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 10637336 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9680995 # MOTIF: [JP] MA0096.1 bZIP910 # MOTIF: [JP] MA0097.1 bZIP911 ---------------------------------------------- Plant J. 1998 Feb;13(4):489-505. Two bZIP proteins from Antirrhinum flowers preferentially bind a hybrid C-box/G-box motif and help to define a new sub-family of bZIP transcription factors. Martinez-Garcia JF, Moyano E, Alcocer MJ, Martin C. Department of Genetics, John Innes Centre, Norwich, UK. Two genes encoding bZIP proteins are expressed in flowers of Antirrhinum majus, predominantly in vascular tissues, carpels and anthers. The sequences of cDNA clones encoding these proteins show them to belong to a distinct sub-family of bZIP proteins which also includes LIP19 from rice and MLIPI5 and OBF1 from maize. The sub-family is characterized by the inclusion of very small proteins consisting of essentially a basic domain and a long leucine zipper. Members also have a conserved upstream open reading frame (uORF) in their 5' leader sequences, implying a common mode of post-transcriptional control. In vitro, the Antirrinum bZIP proteins preferentially bind to a novel hybrid C-box/G-box motif which is found in the promoters of some plant histone genes and of some nuclear-encoded genes with plastidial protein products. Expression of the bZIP proteins in transgenic tobacco under control of the CaMV 35S promoter supports the view that they can regulate expression of genes which contain the preferred target motif within their regulatory sequences, although both enhancement and repression of transcript levels of target genes were observed, indicating that the bZIP proteins probably interact with other factors to modulate transcription in different ways, as has been observed for the small MAF family of bZIP proteins in vertebrates. Publication Types: Research Support, Non-U.S. Gov't PMID: 9680995 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1491700 # MOTIF: [JP] MA0007.1 Ar ---------------------------------------------- Mol Endocrinol. 1992 Dec;6(12):2229-35. A consensus DNA-binding site for the androgen receptor. Roche PJ, Hoare SA, Parker MG. Molecular Endocrinology Laboratory, Imperial Cancer Research Fund, London, United Kingdom. We have used a DNA-binding site selection assay to determine a consensus binding sequence for the androgen receptor (AR). A purified fusion protein containing the AR DNA-binding domain was incubated with a pool of random sequence oligonucleotides, and complexes were isolated by gel mobility shift assays. Individually selected sites were characterised by nucleotide sequencing and compiled to give a consensus AR-binding element. This sequence is comprised of two 6-basepair (bp) asymmetrical elements separated by a 3-bp spacer, 5'-GGA/TACANNNTGTTCT-3', similar to that described for the glucocorticoid response element. Inspection of the consensus revealed a slight preference for G or A nucleotides at the +1 position in the spacer and for A and T nucleotides in the 3'-flanking region. Therefore, a series of oligonucleotides was designed in which the spacer and flanking nucleotides were changed to the least preferred sequence. Competition experiments with these oligonucleotides and the AR fusion protein indicated that an oligonucleotide with both the spacer and flanking sequences changed had greater than 3-fold less affinity than the consensus sequence. The functional activity of these oligonucleotides was also assessed by placing them up-stream of a reporter gene in a transient transfection assay and correlated with the affinity with which the AR fusion protein bound to DNA. Therefore, sequences surrounding the two 6-bp half-sites influence both the binding affinity for the receptor and the functional activity of the response element. Publication Types: Research Support, Non-U.S. Gov't PMID: 1491700 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8132558 # MOTIF: [JP] MA0067.1 Pax2 # MOTIF: [JP] MA0069.1 Pax6 ---------------------------------------------- J Biol Chem. 1994 Mar 18;269(11):8355-61. Identification of a Pax paired domain recognition sequence and evidence for DNA-dependent conformational changes. Epstein J, Cai J, Glaser T, Jepeal L, Maas R. Division of Cardiology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115. Pax genes encode a family of developmentally regulated transcription factors that have been implicated in a number of human and murine congenital disorders, as well as in tumorigenesis (Gruss, P., and Walther, C. (1992) Cell 69, 719-722; Hill, R., and van Heyningen, V. (1992) Trends Genet. 8, 119-120; Chalepakis, G., Tremblay, P., and Gruss, P. (1992) J. Cell Sci. Suppl. 16, 61-67; Maulbecker, C. C., and Gruss, P. (1993) EMBO J. 12, 2361-2367; Walther, C., Guenet, J. L., Simon, D., Deutsch, U., Jostes, B., Goulding, M. D., Plachov, D., Balling, R., and Gruss, P. (1991) Genomics 11, 424-434; Barr, R. G., Galili, N., Holick, J., Biegel, J. A., Rovera, G., and Emanuel, B. S. (1993) Nature Genet. 3, 113-117). These genes are defined by the presence of an evolutionarily conserved DNA binding domain, termed the paired domain. The structure and the DNA binding characteristics of the paired domain remain largely unknown. We have utilized repetitive rounds of a polymerase chain reaction-based selection method to identify the optimal DNA binding sequences for the Pax-2 and Pax-6 paired domains. The results suggest that the paired domain family of peptides bind similar DNA sequences. Identification of this binding site has revealed an important structural clue regarding the mechanism of paired domain binding to DNA. CD and NMR structural analyses of the purified Pax-6 paired domain reveal it to be largely structureless in solution. Upon binding the recognition sequence, the complex becomes markedly less soluble and displays CD spectroscopic evidence of significant alpha-helical structure. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8132558 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8321231 # MOTIF: [JP] MA0029.1 Evi1 ---------------------------------------------- Mol Cell Biol. 1993 Jul;13(7):4291-300. Four of the seven zinc fingers of the Evi-1 myeloid-transforming gene are required for sequence-specific binding to GA(C/T)AAGA(T/C)AAGATAA. Delwel R, Funabiki T, Kreider BL, Morishita K, Ihle JN. Department of Biochemistry, St. Jude Children's Research Hospital, Memphis, Tennessee 38105. Expression of the Evi-1 gene is activated in murine myeloid leukemias by retroviral insertions and in human acute myelogenous leukemia by translocations and inversions involving chromosome band 3q26 where the gene resides. Aberrant expression of the Evi-1 gene has been shown to interfere with myeloid differentiation, which is proposed to be the basis for its role in leukemias. The Evi-1 gene encodes a 145-kDa DNA-binding protein containing two domains of seven and three Cys2-His2 zinc fingers. Previous studies identified a portion of the consensus DNA-binding sequence for the first domain of zinc fingers. The experiments presented here extend these studies and demonstrate that the first domain recognizes a consensus of 15 nucleotides consisting of GA(C/T)AAGA(T/C)AAGATAA. The first three fingers of the first domain do not detectably bind DNA but contribute to the binding by conferring a relative specificity for GACAA verses GATAA in the first position. The first three fingers also contribute to optimal binding of the 15-nucleotide consensus sequence. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8321231 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19339991 # MOTIF: [JP] MA0112.2 ESR1 ---------------------------------------------- EMBO J. 2009 May 20;28(10):1418-28. Epub 2009 Apr 4. ChIP-Seq of ERalpha and RNA polymerase II defines genes differentially responding to ligands. Welboren WJ, van Driel MA, Janssen-Megens EM, van Heeringen SJ, Sweep FC, Span PN, Stunnenberg HG. Department of Molecular Biology, Faculty of Science, Nijmegen Centre for Molecular Life Sciences, Radboud University, Nijmegen, The Netherlands. We used ChIP-Seq to map ERalpha-binding sites and to profile changes in RNA polymerase II (RNAPII) occupancy in MCF-7 cells in response to estradiol (E2), tamoxifen or fulvestrant. We identify 10 205 high confidence ERalpha-binding sites in response to E2 of which 68% contain an estrogen response element (ERE) and only 7% contain a FOXA1 motif. Remarkably, 596 genes change significantly in RNAPII occupancy (59% up and 41% down) already after 1 h of E2 exposure. Although promoter proximal enrichment of RNAPII (PPEP) occurs frequently in MCF-7 cells (17%), it is only observed on a minority of E2-regulated genes (4%). Tamoxifen and fulvestrant partially reduce ERalpha DNA binding and prevent RNAPII loading on the promoter and coding body on E2-upregulated genes. Both ligands act differently on E2-downregulated genes: tamoxifen acts as an agonist thus downregulating these genes, whereas fulvestrant antagonizes E2-induced repression and often increases RNAPII occupancy. Furthermore, our data identify genes preferentially regulated by tamoxifen but not by E2 or fulvestrant. Thus (partial) antagonist loaded ERalpha acts mechanistically different on E2-activated and E2-repressed genes. Publication Types: Research Support, Non-U.S. Gov't PMID: 19339991 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17130146 # MOTIF: [JP] MA0279.1 CAD1 # MOTIF: [JP] MA0287.1 CUP2 # MOTIF: [JP] MA0313.1 HAP2 # MOTIF: [JP] MA0314.1 HAP3 # MOTIF: [JP] MA0316.1 HAP5 # MOTIF: [JP] MA0326.1 MAC1 # MOTIF: [JP] MA0330.1 MBP1::SWI6 # MOTIF: [JP] MA0331.1 MCM1 # MOTIF: [JP] MA0382.1 SKO1 # MOTIF: [JP] MA0384.1 SNT2 # MOTIF: [JP] MA0393.1 STE12 # MOTIF: [JP] MA0407.1 THI2 # MOTIF: [JP] MA0412.1 UME6 # MOTIF: [JP] MA0419.1 YAP7 # MOTIF: [JP] MA0421.1 YDR026C ---------------------------------------------- Nucleic Acids Res. 2007 Jan;35(Database issue):D127-31. Epub 2006 Nov 27. SwissRegulon: a database of genome-wide annotations of regulatory sites. Pachkov M, Erb I, Molina N, van Nimwegen E. Biozentrum, The University of Basel, Klingelbergstrasse 50/70, 4056-CH, Basel, Switzerland. SwissRegulon (http://www.swissregulon.unibas.ch) is a database containing genome-wide annotations of regulatory sites in the intergenic regions of genomes. The regulatory site annotations are produced using a number of recently developed algorithms that operate on multiple alignments of orthologous intergenic regions from related genomes in combination with, whenever available, known sites from the literature, and ChIP-on-chip binding data. Currently SwissRegulon contains annotations for yeast and 17 prokaryotic genomes. The database provides information about the sequence, location, orientation, posterior probability and, whenever available, binding factor of each annotated site. To enable easy viewing of the regulatory site annotations in the context of other features annotated on the genomes, the sites are displayed using the GBrowse genome browser interface and can be queried based on any annotated genomic feature. The database can also be queried for regulons, i.e. sites bound by a common factor. Publication Types: Research Support, Non-U.S. Gov't PMID: 17130146 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15735639 # MOTIF: [JP] PF0001.1 RCGCANGCGY # MOTIF: [JP] PF0002.1 CACGTG # MOTIF: [JP] PF0003.1 SCGGAAGY # MOTIF: [JP] PF0004.1 ACTAYRNNNCCCR # MOTIF: [JP] PF0005.1 GATTGGY # MOTIF: [JP] PF0006.1 GGGCGGR # MOTIF: [JP] PF0007.1 TGANTCA # MOTIF: [JP] PF0008.1 TMTCGCGANR # MOTIF: [JP] PF0009.1 TGAYRTCA # MOTIF: [JP] PF0010.1 GCCATNTTG # MOTIF: [JP] PF0011.1 MGGAAGTG # MOTIF: [JP] PF0012.1 CAGGTG # MOTIF: [JP] PF0013.1 CTTTGT # MOTIF: [JP] PF0014.1 TGACGTCA # MOTIF: [JP] PF0015.1 CAGCTG # MOTIF: [JP] PF0016.1 RYTTCCTG # MOTIF: [JP] PF0017.1 AACTTT # MOTIF: [JP] PF0018.1 TCANNTGAY # MOTIF: [JP] PF0019.1 GKCGCNNNNNNNTGAYG # MOTIF: [JP] PF0020.1 GTGACGY # MOTIF: [JP] PF0021.1 GGAANCGGAANY # MOTIF: [JP] PF0022.1 TGCGCANK # MOTIF: [JP] PF0023.1 TAATTA # MOTIF: [JP] PF0024.1 GGGAGGRR # MOTIF: [JP] PF0025.1 TGACCTY # MOTIF: [JP] PF0026.1 TTAYRTAA # MOTIF: [JP] PF0027.1 TGGNNNNNNKCCAR # MOTIF: [JP] PF0028.1 CTAWWWATA # MOTIF: [JP] PF0029.1 CTTTAAR # MOTIF: [JP] PF0030.1 YGCGYRCGC # MOTIF: [JP] PF0031.1 GGGYGTGNY # MOTIF: [JP] PF0032.1 TGASTMAGC # MOTIF: [JP] PF0033.1 YTATTTTNR # MOTIF: [JP] PF0034.1 CYTAGCAAY # MOTIF: [JP] PF0035.1 GCANCTGNY # MOTIF: [JP] PF0036.1 RTAAACA # MOTIF: [JP] PF0037.1 GTTRYCATRR # MOTIF: [JP] PF0038.1 TGACCTTG # MOTIF: [JP] PF0039.1 TCCCRNNRTGC # MOTIF: [JP] PF0040.1 TTCYNRGAA # MOTIF: [JP] PF0041.1 TGACAGNY # MOTIF: [JP] PF0042.1 TGACATY # MOTIF: [JP] PF0043.1 GTTGNYNNRGNAAC # MOTIF: [JP] PF0044.1 YATGNWAAT # MOTIF: [JP] PF0045.1 CCANNAGRKGGC # MOTIF: [JP] PF0046.1 WTTGKCTG # MOTIF: [JP] PF0047.1 TGCCAAR # MOTIF: [JP] PF0048.1 GCGNNANTTCC # MOTIF: [JP] PF0049.1 CATTGTYY # MOTIF: [JP] PF0050.1 RGAGGAARY # MOTIF: [JP] PF0051.1 TATAAA # MOTIF: [JP] PF0052.1 YYCATTCAWW # MOTIF: [JP] PF0053.1 RYTGCNNRGNAAC # MOTIF: [JP] PF0054.1 TAAWWATAG # MOTIF: [JP] PF0055.1 TGGAAA # MOTIF: [JP] PF0056.1 GGGTGGRR # MOTIF: [JP] PF0057.1 ACCTGTTG # MOTIF: [JP] PF0058.1 YCATTAA # MOTIF: [JP] PF0059.1 WCTCNATGGY # MOTIF: [JP] PF0060.1 TTGTTT # MOTIF: [JP] PF0061.1 YTAATTAA # MOTIF: [JP] PF0062.1 SMTTTTGT # MOTIF: [JP] PF0063.1 AAGWWRNYGGC # MOTIF: [JP] PF0064.1 TTANTCA # MOTIF: [JP] PF0065.1 ARGGGTTAA # MOTIF: [JP] PF0066.1 RACTNNRTTTNC # MOTIF: [JP] PF0067.1 TGANNYRGCA # MOTIF: [JP] PF0068.1 RGAANNTTC # MOTIF: [JP] PF0069.1 SGCGSSAAA # MOTIF: [JP] PF0070.1 CGTSACG # MOTIF: [JP] PF0071.1 SYATTGTG # MOTIF: [JP] PF0072.1 TTCYRGAA # MOTIF: [JP] PF0073.1 CTTTGA # MOTIF: [JP] PF0074.1 GGAMTNNNNNTCCY # MOTIF: [JP] PF0075.1 TNCATNTCCYR # MOTIF: [JP] PF0076.1 CAGGTA # MOTIF: [JP] PF0077.1 AAAYRNCTG # MOTIF: [JP] PF0078.1 GCTNWTTGK # MOTIF: [JP] PF0079.1 WGGAATGY # MOTIF: [JP] PF0080.1 SNACANNNYSYAGA # MOTIF: [JP] PF0081.1 CGGAARNGGCNG # MOTIF: [JP] PF0082.1 CTGYNNCTYTAA # MOTIF: [JP] PF0083.1 TGTTTGY # MOTIF: [JP] PF0084.1 RGTTAMWNATT # MOTIF: [JP] PF0085.1 STTTCRNTTT # MOTIF: [JP] PF0086.1 GGGNNTTTCC # MOTIF: [JP] PF0087.1 RYTGCNWTGGNR # MOTIF: [JP] PF0088.1 GGCNKCCATNK # MOTIF: [JP] PF0089.1 GTTNYYNNGGTNA # MOTIF: [JP] PF0090.1 YAATNRNNNYNATT # MOTIF: [JP] PF0091.1 GTGGGTGK # MOTIF: [JP] PF0092.1 TGCTGAY # MOTIF: [JP] PF0093.1 GGATTA # MOTIF: [JP] PF0094.1 TGATTTRY # MOTIF: [JP] PF0095.1 GCCNNNWTAAR # MOTIF: [JP] PF0096.1 YGCANTGCR # MOTIF: [JP] PF0097.1 YATTNATC # MOTIF: [JP] PF0098.1 GTCNYYATGR # MOTIF: [JP] PF0099.1 ATCMNTCCGY # MOTIF: [JP] PF0100.1 CRGAARNNNNCGA # MOTIF: [JP] PF0101.1 CTGCAGY # MOTIF: [JP] PF0102.1 ATGGYGGA # MOTIF: [JP] PF0103.1 ACAWNRNSRCGG # MOTIF: [JP] PF0104.1 CCAATNNSNNNGCG # MOTIF: [JP] PF0105.1 ACTWSNACTNY # MOTIF: [JP] PF0106.1 CCGNMNNTNACG # MOTIF: [JP] PF0107.1 RTTTNNNYTGGM # MOTIF: [JP] PF0108.1 AACWWCAANK # MOTIF: [JP] PF0109.1 YGTCCTTGR # MOTIF: [JP] PF0110.1 MCAATNNNNNGCG # MOTIF: [JP] PF0111.1 RACCACAR # MOTIF: [JP] PF0112.1 KTGGYRSGAA # MOTIF: [JP] PF0113.1 AACYNNNNTTCCS # MOTIF: [JP] PF0114.1 YTCCCRNNAGGY # MOTIF: [JP] PF0115.1 YRTCANNRCGC # MOTIF: [JP] PF0116.1 KMCATNNWGGA # MOTIF: [JP] PF0117.1 TGTYNNNNNRGCARM # MOTIF: [JP] PF0118.1 GGCNRNWCTTYS # MOTIF: [JP] PF0119.1 GGGNRMNNYCAT # MOTIF: [JP] PF0120.1 KRCTCNNNNMANAGC # MOTIF: [JP] PF0121.1 CCAWWNAAGG # MOTIF: [JP] PF0122.1 RNTCANNRNNYNATTW # MOTIF: [JP] PF0123.1 GGCNNMSMYNTTG # MOTIF: [JP] PF0124.1 CCAWYNNGAAR # MOTIF: [JP] PF0125.1 RAAGNYNNCTTY # MOTIF: [JP] PF0126.1 WYAAANNRNNNGCG # MOTIF: [JP] PF0127.1 WWTAAGGC # MOTIF: [JP] PF0128.1 RYCACNNRNNRNCAG # MOTIF: [JP] PF0129.1 RRAGTTGT # MOTIF: [JP] PF0130.1 CCCNNGGGAR # MOTIF: [JP] PF0131.1 GATAAGR # MOTIF: [JP] PF0132.1 TCCATTKW # MOTIF: [JP] PF0133.1 RYTAAWNNNTGAY # MOTIF: [JP] PF0134.1 CATRRAGC # MOTIF: [JP] PF0135.1 AGCYRWTTC # MOTIF: [JP] PF0136.1 TAAYNRNNTCC # MOTIF: [JP] PF0137.1 GAANYNYGACNY # MOTIF: [JP] PF0138.1 MYAATNNNNNNNGGC # MOTIF: [JP] PF0139.1 AAAYWAACM # MOTIF: [JP] PF0140.1 RNGTGGGC # MOTIF: [JP] PF0141.1 TTCNRGNNNNTTC # MOTIF: [JP] PF0142.1 ACAWYAAAG # MOTIF: [JP] PF0143.1 CAGNWMCNNNGAC # MOTIF: [JP] PF0144.1 AAANWWTGC # MOTIF: [JP] PF0145.1 YKACATTT # MOTIF: [JP] PF0146.1 RRCCGTTA # MOTIF: [JP] PF0147.1 YAATNANRNNNCAG # MOTIF: [JP] PF0148.1 GATGKMRGCG # MOTIF: [JP] PF0149.1 YGACNNYACAR # MOTIF: [JP] PF0150.1 YTTCCNNNGGAMR # MOTIF: [JP] PF0151.1 RYAAAKNNNNNNTTGW # MOTIF: [JP] PF0152.1 WCAANNNYCAG # MOTIF: [JP] PF0153.1 CTGRYYYNATT # MOTIF: [JP] PF0154.1 RNCTGNYNRNCTGNY # MOTIF: [JP] PF0155.1 WGTTNNNNNAAA # MOTIF: [JP] PF0156.1 YRCCAKNNGNCGC # MOTIF: [JP] PF0157.1 KCCGNSWTTT # MOTIF: [JP] PF0158.1 CCCNNNNNNAAGWT # MOTIF: [JP] PF0159.1 GGCKCATGS # MOTIF: [JP] PF0160.1 CAGNYGKNAAA # MOTIF: [JP] PF0161.1 TTANWNANTGGM # MOTIF: [JP] PF0162.1 TAANNYSGCG # MOTIF: [JP] PF0163.1 GGARNTKYCCA # MOTIF: [JP] PF0164.1 GCGSCMNTTT # MOTIF: [JP] PF0165.1 CCAWNWWNNNGGC # MOTIF: [JP] PF0166.1 YNTTTNNNANGCARM # MOTIF: [JP] PF0167.1 CCTNTMAGA # MOTIF: [JP] PF0168.1 YTAAYNGCT # MOTIF: [JP] PF0169.1 TTTNNANAGCYR # MOTIF: [JP] PF0170.1 YNGTTNNNATT # MOTIF: [JP] PF0171.1 CTCNANGTGNY # MOTIF: [JP] PF0172.1 TTGCWCAAY # MOTIF: [JP] PF0173.1 YWATTWNNRGCT # MOTIF: [JP] PF0174.1 WTGAAAT ---------------------------------------------- Nature. 2005 Mar 17;434(7031):338-45. Epub 2005 Feb 27. Systematic discovery of regulatory motifs in human promoters and 3' UTRs by comparison of several mammals. Xie X, Lu J, Kulbokas EJ, Golub TR, Mootha V, Lindblad-Toh K, Lander ES, Kellis M. Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02141, USA. Comprehensive identification of all functional elements encoded in the human genome is a fundamental need in biomedical research. Here, we present a comparative analysis of the human, mouse, rat and dog genomes to create a systematic catalogue of common regulatory motifs in promoters and 3' untranslated regions (3' UTRs). The promoter analysis yields 174 candidate motifs, including most previously known transcription-factor binding sites and 105 new motifs. The 3'-UTR analysis yields 106 motifs likely to be involved in post-transcriptional regulation. Nearly one-half are associated with microRNAs (miRNAs), leading to the discovery of many new miRNA genes and their likely target genes. Our results suggest that previous estimates of the number of human miRNA genes were low, and that miRNAs regulate at least 20% of human genes. The overall results provide a systematic view of gene regulation in the human, which will be refined as additional mammalian genomes become available. Publication Types: Comparative Study Research Support, U.S. Gov't, P.H.S. PMID: 15735639 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17606643 # MOTIF: [JP] MA0260.1 che-1 ---------------------------------------------- Genes Dev. 2007 Jul 1;21(13):1653-74. The molecular signature and cis-regulatory architecture of a C. elegans gustatory neuron. Etchberger JF, Lorch A, Sleumer MC, Zapf R, Jones SJ, Marra MA, Holt RA, Moerman DG, Hobert O. Howard Hughes Medical Institute, Department of Biochemistry and Molecular Biophysics, Columbia University Medical Center, New York, NY 10032, USA. Taste receptor cells constitute a highly specialized cell type that perceives and conveys specific sensory information to the brain. The detailed molecular composition of these cells and the mechanisms that program their fate are, in general, poorly understood. We have generated serial analysis of gene expression (SAGE) libraries from two distinct populations of single, isolated sensory neuron classes, the gustatory neuron class ASE and the thermosensory neuron class AFD, from the nematode Caenorhabditis elegans. By comparing these two libraries, we have identified >1000 genes that define the ASE gustatory neuron class on a molecular level. This set of genes contains determinants of the differentiated state of the ASE neuron, such as a surprisingly complex repertoire of transcription factors (TFs), ion channels, neurotransmitters, and receptors, as well as seven-transmembrane receptor (7TMR)-type putative gustatory receptor genes. Through the in vivo dissection of the cis-regulatory regions of several ASE-expressed genes, we identified a small cis-regulatory motif, the "ASE motif," that is required for the expression of many ASE-expressed genes. We demonstrate that the ASE motif is a binding site for the C2H2 zinc finger TF CHE-1, which is essential for the correct differentiation of the ASE gustatory neuron. Taken together, our results provide a unique view of the molecular landscape of a single neuron type and reveal an important aspect of the regulatory logic for gustatory neuron specification in C. elegans. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 17606643 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12198246 # MOTIF: [JP] MA0109.1 Hltf ---------------------------------------------- Mol Endocrinol. 2002 Sep;16(9):2101-12. Identification of the RUSH consensus-binding site by cyclic amplification and selection of targets: demonstration that RUSH mediates the ability of prolactin to augment progesterone-dependent gene expression. Hewetson A, Hendrix EC, Mansharamani M, Lee VH, Chilton BS. Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, Texas 79430, USA. RUSH-1alpha(beta) transcription factors were cloned by recognition site screening with an 85-bp region (-170/-85) of the rabbit uteroglobin gene. Deletion analysis showed this region was essential to prolactin (PRL) action, but conclusions were limited by the complexity of the large deletion. Cyclic amplification and selection of targets (CASTing) was used to identify the RUSH-binding site (-126/-121). Endometrial nuclear proteins were incubated with a pool of degenerate oligonucleotides and immunoprecipitated with RUSH-1alpha(beta) antibodies. Bound DNA was amplified by PCR. The consensus motif (MCWTDK) was identified after five rounds of CASTing, authenticated by CASTing with RUSH-1alpha-specific antibodies and recombinant protein, and refined with EMSA. Dissociation rate constants (K(d) = 0.1-1.0 nM; r = 0.99) revealed high-affinity binding. Chromatin immunoprecipitation confirmed in vivo binding of RUSH to the transcriptionally active uteroglobin promoter. CASTing also revealed RUSH-GATA transcription factor interactions. Endometrial GATA-4 expression is progesterone dependent (Northern analysis) and preferentially localized in the epithelium (in situ hybridization). Although physically affiliated with RUSH, uterine forms of GATA-4 were not required for RUSH-DNA binding. Site-directed mutagenesis and transient transfection assays showed the RUSH motif mediates the ability of PRL to augment progesterone-dependent uteroglobin transcription. RUSH is central to the mechanism whereby PRL augments progesterone-dependent gene transcription. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 12198246 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15572468 # MOTIF: [JP] MA0185.1 Deaf1 # MOTIF: [JP] MA0205.1 Trl # MOTIF: [JP] MA0213.1 brk # MOTIF: [JP] MA0237.1 pan # MOTIF: [JP] MA0243.1 sd # MOTIF: [JP] MA0244.1 slbo # MOTIF: [JP] MA0249.1 twi # MOTIF: [JP] MA0254.1 vvl # MOTIF: [JP] MA0255.1 z ---------------------------------------------- Bioinformatics. 2005 Apr 15;21(8):1747-9. Epub 2004 Nov 30. Drosophila DNase I footprint database: a systematic genome annotation of transcription factor binding sites in the fruitfly, Drosophila melanogaster. Bergman CM, Carlson JW, Celniker SE. Department of Genetics, University of Cambridge, Cambridge CB2 3EH, UK. cbergman@gen.cam.ac.uk SUMMARY: Despite increasing numbers of computational tools developed to predict cis-regulatory sequences, the availability of high-quality datasets of transcription factor binding sites limits advances in the bioinformatics of gene regulation. Here we present such a dataset based on a systematic literature curation and genome annotation of DNase I footprints for the fruitfly, Drosophila melanogaster. Using the experimental results of 201 primary references, we annotated 1367 binding sites from 87 transcription factors and 101 target genes in the D.melanogaster genome sequence. These data will provide a rich resource for future bioinformatics analyses of transcriptional regulation in Drosophila such as constructing motif models, training cis-regulatory module detectors, benchmarking alignment tools and continued text mining of the extensive literature on transcriptional regulation in this important model organism. AVAILABILITY: http://www.flyreg.org/ CONTACT: cbergman@gen.cam.ac.uk. Publication Types: Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S. PMID: 15572468 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 11247607 # MOTIF: [JP] MA0110.1 ATHB-5 ---------------------------------------------- Plant Mol Biol. 2001 Jan;45(1):63-73. DNA-binding and dimerization preferences of Arabidopsis homeodomain-leucine zipper transcription factors in vitro. Johannesson H, Wang Y, Engstrom P. Department of Evolutionary Biology, Physiological Botany, Uppsala, Sweden. Homeodomain-leucine zipper (HDZip) proteins constitute a large family of transcription factors apparently unique to plants. In this report we characterize the DNA-binding and dimerization preferences in vitro of class I HDZip proteins. Using gel-exclusion chromatography and in vitro protein binding assays we demonstrate that the HDZip class I protein ATHB5 forms a homodimeric complex in solution. Consistent with this finding we have demonstrated the sequence-specific interaction of ATHB5 with a 9 bp pseudopalindromic DNA sequence, CAATNATTG, composed of two half-sites overlapping at a central position, by use of a PCR-assisted binding-site selection assay and competitive EMSA experiments. A majority of other known members of HDZip class I interacted with similar DNA sequences, but differed in their preference for A/T versus G/C in the central position of the binding site. Selective heterodimerization in vitro was demonstrated between ATHB5 and different class I HDZip proteins. Heterodimer formation between class I HDZip proteins is of potential functional significance for the integration of information from different signalling pathways in the control of plant development. Publication Types: Research Support, Non-U.S. Gov't PMID: 11247607 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18272478 # MOTIF: [JP] MA0258.1 ESR2 ---------------------------------------------- Proc Natl Acad Sci U S A. 2008 Feb 19;105(7):2604-9. Epub 2008 Feb 13. The genome landscape of ERalpha- and ERbeta-binding DNA regions. Liu Y, Gao H, Marstrand TT, Strom A, Valen E, Sandelin A, Gustafsson JA, Dahlman-Wright K. Department of Biosciences and Nutrition, Karolinska Institutet, Novum, SE-14157 Huddinge, Sweden. In this article, we have applied the ChIP-on-chip approach to pursue a large scale identification of ERalpha- and ERbeta-binding DNA regions in intact chromatin. We show that there is a high degree of overlap between the regions identified as bound by ERalpha and ERbeta, respectively, but there are also regions that are bound by ERalpha only in the presence of ERbeta, as well as regions that are selectively bound by either receptor. Analysis of bound regions shows that regions bound by ERalpha have distinct properties in terms of genome landscape, sequence features, and conservation compared with regions that are bound by ERbeta. ERbeta-bound regions are, as a group, located more closely to transcription start sites. ERalpha- and ERbeta-bound regions differ in sequence properties, with ERalpha-bound regions having an overrepresentation of TA-rich motifs including forkhead binding sites and ERbeta-bound regions having a predominance of classical estrogen response elements (EREs) and GC-rich motifs. Differences in the properties of ER bound regions might explain some of the differences in gene expression programs and physiological effects shown by the respective estrogen receptors. Publication Types: Research Support, Non-U.S. Gov't PMID: 18272478 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10848601 # MOTIF: [JP] POL005.1 DPE ---------------------------------------------- Mol Cell Biol. 2000 Jul;20(13):4754-64. The downstream promoter element DPE appears to be as widely used as the TATA box in Drosophila core promoters. Kutach AK, Kadonaga JT. Department of Biology and Center for Molecular Genetics, University of California, San Diego, La Jolla 92093-0347, USA. The downstream promoter element (DPE) functions cooperatively with the initiator (Inr) for the binding of TFIID in the transcription of core promoters in the absence of a TATA box. We examined the properties of sequences that can function as a DPE as well as the range of promoters that use the DPE as a core promoter element. By using an in vitro transcription assay, we identified 17 new DPE-dependent promoters and found that all possessed identical spacing between the Inr and DPE. Moreover, mutational analysis indicated that the insertion or deletion of a single nucleotide between the Inr and DPE causes a reduction in transcriptional activity and TFIID binding. To explore the range of sequences that can function as a DPE, we constructed and analyzed randomized promoter libraries. These experiments yielded the DPE functional range set, which represents sequences that contribute to or are compatible with DPE function. We then analyzed the DPE functional range set in conjunction with a Drosophila core promoter database that we compiled from 205 promoters with accurately mapped start sites. Somewhat surprisingly, the DPE sequence motif is as common as the TATA box in Drosophila promoters. There is, in addition, a striking adherence of Inr sequences to the Inr consensus in DPE-containing promoters relative to DPE-less promoters. Furthermore, statistical and biochemical analyses indicated that a G nucleotide between the Inr and DPE contributes to transcription from DPE-containing promoters. Thus, these data reveal that the DPE exhibits a strict spacing requirement yet some sequence flexibility and appears to be as widely used as the TATA box in Drosophila. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 10848601 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7791788 # MOTIF: [JP] MA0092.1 Hand1::Tcfe2a ---------------------------------------------- Mol Cell Biol. 1995 Jul;15(7):3813-22. Identification of a new family of tissue-specific basic helix-loop-helix proteins with a two-hybrid system. Hollenberg SM, Sternglanz R, Cheng PF, Weintraub H. Howard Hughes Medical Institute, Fred Hutchinson Cancer Research Center, Seattle, Washington 98104, USA. With modified two-hybrid technology, we have isolated a member of a new family of basic helix-loop-helix (bHLH) transcription factors. Thing1 (Th1) was identified in a screen of a mouse embryo cDNA library as a partner for the Drosophila E protein daughterless. RNA in situ hybridization and reverse transcriptase-PCR demonstrate a stage- and tissue-specific distribution for the expression of Th1. Although tissue specific, the expression pattern of Th1 is fairly complex. During development, Th1 mRNA is widely expressed in extraembryonic tissues, portions of the heart, autonomic ganglia, the gut, and pharyngeal arches. At embryonic day 7.5 (E7.5), extraembryonic derivatives show robust Th1 expression. By E8.5, expression in the embryonic heart becomes detectable. During the next 2 days of development, the signal also includes gut and pharyngeal arches. Predominant expression at E13.5 is in neural crest derivatives, especially the autonomic nervous system and adrenal medulla. Expression of Th1 persists in the adult, in which it is localized to the smooth muscle cells of the gut. In vitro, Th1 protein recognizes a set of DNA sites that are more degenerate than has been determined for other bHLH factors, indicating a reduced binding specificity. Transient transfection of NIH 3T3 cells with GAL4-Th1 fusions reveals a repression activity mediated by the Th1 bHLH domain. In combination, these properties define Th1 as a new bHLH protein with a unique set of properties. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 7791788 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1594445 # MOTIF: [JP] MA0104.1 Mycn ---------------------------------------------- Nucleic Acids Res. 1992 May 11;20(9):2257-63. Determination of the DNA sequence recognized by the bHLH-zip domain of the N-Myc protein. Alex R, Sozeri O, Meyer S, Dildrop R. Institute for Genetics, University of Cologne, Germany. The DNA-binding domain of the murine N-Myc protein, comprising the basic helix-loop-helix-zipper (bHLH-zip) region was expressed as a fusion protein in E. coli. The affinity purified glutathione-S-transferase-N-Myc fusion protein (GST-N-MYC) was used to select the N-Myc specific DNA-recognition motif from a pool of random-sequence oligonucleotides. After seven rounds of binding-site selection, specifically enriched oligonucleotides were cloned and sequenced. Of 31 individual oligonucleotides whose sequences were determined, 30 contained a common DNA-motif, defining the hexameric consensus sequence CACGTG. We confirm by mutational analysis that binding of the N-Myc derived bHLH-zip domain to this motif is sequence-specific. Publication Types: Research Support, Non-U.S. Gov't PMID: 1594445 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18332042 # MOTIF: [JP] MA00445.1 D # MOTIF: [JP] MA0165.1 Abd-B # MOTIF: [JP] MA0166.1 Antp # MOTIF: [JP] MA0167.1 Awh # MOTIF: [JP] MA0168.1 B-H1 # MOTIF: [JP] MA0169.1 B-H2 # MOTIF: [JP] MA0170.1 C15 # MOTIF: [JP] MA0171.1 CG11085 # MOTIF: [JP] MA0172.1 CG11294 # MOTIF: [JP] MA0173.1 CG11617 # MOTIF: [JP] MA0174.1 CG42234 # MOTIF: [JP] MA0175.1 CG13424 # MOTIF: [JP] MA0176.1 CG15696 # MOTIF: [JP] MA0177.1 CG18599 # MOTIF: [JP] MA0178.1 CG32105 # MOTIF: [JP] MA0179.1 CG32532 # MOTIF: [JP] MA0181.1 Vsx1 # MOTIF: [JP] MA0182.1 CG4328 # MOTIF: [JP] MA0184.1 CG9876 # MOTIF: [JP] MA0186.1 Dfd # MOTIF: [JP] MA0188.1 Dr # MOTIF: [JP] MA0189.1 E5 # MOTIF: [JP] MA0190.1 Gsc # MOTIF: [JP] MA0191.1 HGTX # MOTIF: [JP] MA0192.1 Hmx # MOTIF: [JP] MA0193.1 Lag1 # MOTIF: [JP] MA0194.1 Lim1 # MOTIF: [JP] MA0195.1 Lim3 # MOTIF: [JP] MA0196.1 NK7.1 # MOTIF: [JP] MA0198.1 OdsH # MOTIF: [JP] MA0199.1 Optix # MOTIF: [JP] MA0200.1 Pph13 # MOTIF: [JP] MA0202.1 Rx # MOTIF: [JP] MA0203.1 Scr # MOTIF: [JP] MA0204.1 Six4 # MOTIF: [JP] MA0206.1 abd-A # MOTIF: [JP] MA0207.1 achi # MOTIF: [JP] MA0209.1 ap # MOTIF: [JP] MA0210.1 ara # MOTIF: [JP] MA0211.1 bap # MOTIF: [JP] MA0212.1 bcd # MOTIF: [JP] MA0214.1 bsh # MOTIF: [JP] MA0215.1 btn # MOTIF: [JP] MA0216.1 cad # MOTIF: [JP] MA0217.1 caup # MOTIF: [JP] MA0218.1 ct # MOTIF: [JP] MA0219.1 ems # MOTIF: [JP] MA0220.1 en # MOTIF: [JP] MA0221.1 eve # MOTIF: [JP] MA0223.1 exex # MOTIF: [JP] MA0225.1 ftz # MOTIF: [JP] MA0226.1 hbn # MOTIF: [JP] MA0227.1 hth # MOTIF: [JP] MA0228.1 ind # MOTIF: [JP] MA0230.1 lab # MOTIF: [JP] MA0231.1 lbe # MOTIF: [JP] MA0232.1 lbl # MOTIF: [JP] MA0233.1 mirr # MOTIF: [JP] MA0234.1 oc # MOTIF: [JP] MA0235.1 onecut # MOTIF: [JP] MA0236.1 otp # MOTIF: [JP] MA0238.1 pb # MOTIF: [JP] MA0240.1 repo # MOTIF: [JP] MA0241.1 ro # MOTIF: [JP] MA0245.1 slou # MOTIF: [JP] MA0246.1 so # MOTIF: [JP] MA0248.1 tup # MOTIF: [JP] MA0250.1 unc-4 # MOTIF: [JP] MA0251.1 unpg # MOTIF: [JP] MA0252.1 vis # MOTIF: [JP] MA0256.1 zen # MOTIF: [JP] MA0257.1 zen2 # MOTIF: [JP] MA0443.1 btd # MOTIF: [JP] MA0444.1 CG34031 # MOTIF: [JP] MA0446.1 fkh # MOTIF: [JP] MA0447.1 gt # MOTIF: [JP] MA0448.1 H2.0 # MOTIF: [JP] MA0449.1 h # MOTIF: [JP] MA0450.1 hkb # MOTIF: [JP] MA0451.1 kni # MOTIF: [JP] MA0452.1 Kr # MOTIF: [JP] MA0453.1 nub # MOTIF: [JP] MA0454.1 odd # MOTIF: [JP] MA0455.1 OdsH # MOTIF: [JP] MA0456.1 opa # MOTIF: [JP] MA0457.1 PHDP # MOTIF: [JP] MA0458.1 slp1 # MOTIF: [JP] MA0459.1 tll # MOTIF: [JP] MA0460.1 ttk ---------------------------------------------- Nucleic Acids Res. 2008 May;36(8):2547-60. Epub 2008 Mar 10. A systematic characterization of factors that regulate Drosophila segmentation via a bacterial one-hybrid system. Noyes MB, Meng X, Wakabayashi A, Sinha S, Brodsky MH, Wolfe SA. Program in Gene Function and Expression, Department of Biochemistry and Molecular Pharmacology, Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA. Specificity data for groups of transcription factors (TFs) in a common regulatory network can be used to computationally identify the location of cis-regulatory modules in a genome. The primary limitation for this type of analysis is the paucity of specificity data that is available for the majority of TFs. We describe an omega-based bacterial one-hybrid system that provides a rapid method for characterizing DNA-binding specificities on a genome-wide scale. Using this system, 35 members of the Drosophila melanogaster segmentation network have been characterized, including representative members of all of the major classes of DNA-binding domains. A suite of web-based tools was created that uses this binding site dataset and phylogenetic comparisons to identify cis-regulatory modules throughout the fly genome. These tools allow specificities for any combination of factors to be used to perform rapid local or genome-wide searches for cis-regulatory modules. The utility of these factor specificities and tools is demonstrated on the well-characterized segmentation network. By incorporating specificity data on an additional 66 factors that we have characterized, our tools utilize approximately 14% of the predicted factors within the fly genome and provide an important new community resource for the identification of cis-regulatory modules. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 18332042 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1542566 # MOTIF: [JP] MA0098.1 ETS1 ---------------------------------------------- Nucleic Acids Res. 1992 Feb 25;20(4):699-704. Identification of nucleotide preferences in DNA sequences recognised specifically by c-Ets-1 protein. Woods DB, Ghysdael J, Owen MJ. Imperial Cancer Research Fund, London, UK. The protooncogene Ets-1 is a member of the c-Ets family of genes originally identified through their sequence homology to the v-ets gene of the avian erythroblastosis virus E26. Ets-like factors are characterised by a conserved 85 amino acid domain which appears to be essential for binding to purine rich DNA sequences. Sequences binding to Ets-1 were selected from a random oligonucleotide pool by immunoprecipitation and amplified using the Polymerase Chain Reaction. Oligonucleotides enriched by this procedure were cloned in plasmids and sequenced. Alignment of DNA sequences revealed GGAA and GGAT cores at about a 1.4:1 ratio. Preferred sequences were identified both 5' and 3' of the GGAW core, extending the binding site to ACMGGAWRTT. Analysis of the flanking sequences associated with GGAA and GGAT cores revealed differences which may have compensated for the generally lower affinity of binding sites containing a GGAT core. Lastly mutational analysis of one particular Ets-1 binding site was used to establish the relative importance for binding of some nucleotides within the core and to show that Ets-1 and the closely related Ets-2 proteins bind to similar sequences. PMID: 1542566 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17512414 # MOTIF: [JP] MA0139.1 CTCF ---------------------------------------------- Cell. 2007 May 18;129(4):823-37. Comment in: Cell. 2007 Nov 30;131(5):831-2; author reply 832-3. High-resolution profiling of histone methylations in the human genome. Barski A, Cuddapah S, Cui K, Roh TY, Schones DE, Wang Z, Wei G, Chepelev I, Zhao K. Laboratory of Molecular Immunology, National Heart, Lung, and Blood Institute, NIH, Bethesda, MD 20892, USA. Histone modifications are implicated in influencing gene expression. We have generated high-resolution maps for the genome-wide distribution of 20 histone lysine and arginine methylations as well as histone variant H2A.Z, RNA polymerase II, and the insulator binding protein CTCF across the human genome using the Solexa 1G sequencing technology. Typical patterns of histone methylations exhibited at promoters, insulators, enhancers, and transcribed regions are identified. The monomethylations of H3K27, H3K9, H4K20, H3K79, and H2BK5 are all linked to gene activation, whereas trimethylations of H3K27, H3K9, and H3K79 are linked to repression. H2A.Z associates with functional regulatory elements, and CTCF marks boundaries of histone methylation domains. Chromosome banding patterns are correlated with unique patterns of histone modifications. Chromosome breakpoints detected in T cell cancers frequently reside in chromatin regions associated with H3K4 methylations. Our data provide new insights into the function of histone methylation and chromatin organization in genome function. Publication Types: Research Support, N.I.H., Intramural PMID: 17512414 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1290524 # MOTIF: [JP] MA0015.1 Cf2_II ---------------------------------------------- Science. 1992 Sep 25;257(5078):1951-5. Sequence discrimination by alternatively spliced isoforms of a DNA binding zinc finger domain. Gogos JA, Hsu T, Bolton J, Kafatos FC. Department of Cellular and Developmental Biology, Harvard University, Cambridge, MA 02138. Two major developmentally regulated isoforms of the Drosophila chorion transcription factor CF2 differ by an extra zinc finger within the DNA binding domain. The preferred DNA binding sites were determined and are distinguished by an internal duplication of TAT in the site recognized by the isoform with the extra finger. The results are consistent with modular interactions between zinc fingers and trinucleotides and also suggest rules for recognition of AT-rich DNA sites by zinc finger proteins. The results show how modular finger interactions with trinucleotides can be used, in conjunction with alternative splicing, to alter the binding specificity and increase the spectrum of sites recognized by a DNA binding domain. Thus, CF2 may potentially regulate distinct sets of target genes during development. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 1290524 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16522208 # MOTIF: [JP] MA0271.1 ARG80 # MOTIF: [JP] MA0272.1 ARG81 # MOTIF: [JP] MA0274.1 ARR1 # MOTIF: [JP] MA0276.1 ASH1 # MOTIF: [JP] MA0290.1 DAL81 # MOTIF: [JP] MA0305.1 GCR2 # MOTIF: [JP] MA0315.1 HAP4 # MOTIF: [JP] MA0320.1 IME1 # MOTIF: [JP] MA0321.1 INO2 # MOTIF: [JP] MA0322.1 INO4 # MOTIF: [JP] MA0323.1 IXR1 # MOTIF: [JP] MA0327.1 MATA1 # MOTIF: [JP] MA0332.1 MET28 # MOTIF: [JP] MA0335.1 MET4 # MOTIF: [JP] MA0340.1 MOT3 # MOTIF: [JP] MA0349.1 OPI1 # MOTIF: [JP] MA0353.1 PDR3 # MOTIF: [JP] MA0370.1 RME1 # MOTIF: [JP] MA0387.1 SPT2 # MOTIF: [JP] MA0388.1 SPT23 # MOTIF: [JP] MA0394.1 STP1 # MOTIF: [JP] MA0399.1 SUT1 # MOTIF: [JP] MA0417.1 YAP5 # MOTIF: [JP] MA0426.1 YHP1 # MOTIF: [JP] MA0440.1 ZAP1 ---------------------------------------------- BMC Bioinformatics. 2006 Mar 7;7:113. An improved map of conserved regulatory sites for Saccharomyces cerevisiae. MacIsaac KD, Wang T, Gordon DB, Gifford DK, Stormo GD, Fraenkel E. Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, MA 02142, USA. macisaac@mit.edu BACKGROUND: The regulatory map of a genome consists of the binding sites for proteins that determine the transcription of nearby genes. An initial regulatory map for S. cerevisiae was recently published using six motif discovery programs to analyze genome-wide chromatin immunoprecipitation data for 203 transcription factors. The programs were used to identify sequence motifs that were likely to correspond to the DNA-binding specificity of the immunoprecipitated proteins. We report improved versions of two conservation-based motif discovery algorithms, PhyloCon and Converge. Using these programs, we create a refined regulatory map for S. cerevisiae by reanalyzing the same chromatin immunoprecipitation data. RESULTS: Applying the same conservative criteria that were applied in the original study, we find that PhyloCon and Converge each separately discover more known specificities than the combination of all six programs in the previous study. Combining the results of PhyloCon and Converge, we discover significant sequence motifs for 36 transcription factors that were previously missed. The new set of motifs identifies 636 more regulatory interactions than the previous one. The new network contains 28% more regulatory interactions among transcription factors, evidence of greater cross-talk between regulators. CONCLUSION: Combining two complementary computational strategies for conservation-based motif discovery improves the ability to identify the specificity of transcriptional regulators from genome-wide chromatin immunoprecipitation data. The increased sensitivity of these methods significantly expands the map of yeast regulatory sites without the need to alter any of the thresholds for statistical significance. The new map of regulatory sites reveals a more elaborate and complex view of the yeast genetic regulatory network than was observed previously. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 16522208 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16997917 # MOTIF: [JP] MA0125.1 Nobox ---------------------------------------------- J Biol Chem. 2006 Nov 24;281(47):35747-56. Epub 2006 Sep 22. Characterization of NOBOX DNA binding specificity and its regulation of Gdf9 and Pou5f1 promoters. Choi Y, Rajkovic A. Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas 77030, USA. Nobox (newborn ovary homeobox gene) deficiency disrupts early folliculogenesis and the expression of oocyte-specific genes in mice. Here, we identified several cis-acting sites, TAATTG, TAGTTG, and TAATTA as NOBOX DNA binding elements (NBEs) using a library of randomly generated oligonucleotides by cyclic amplification of sequence target assay and mutation analyses. We show that NOBOX preferentially binds to the NOBOX binding elements with high affinity. In addition, we found that promoter regions of mouse Pou5f1 and Gdf9 contain one (-426) and three NOBOX binding elements (-786, -967, and -1259), respectively. NOBOX binds to these putative NOBOX binding elements with high affinity and augmented transcriptional activity of luciferase reporter driven by mouse Pou5f1 and Gdf9 promoters containing the NOBOX binding elements. In chromatin immunoprecipitation assays, DNA sequences from Pou5f1 and Gdf9 promoters co-precipitated with anti-NOBOX antibody. These results suggest that NOBOX directly regulates the transcription of Pou5f1 and Gdf9 in oocytes during early folliculogenesis. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 16997917 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15020707 # MOTIF: [JP] MA0120.1 id1 ---------------------------------------------- Nucleic Acids Res. 2004 Mar 12;32(5):1710-20. Print 2004. The maize ID1 flowering time regulator is a zinc finger protein with novel DNA binding properties. Kozaki A, Hake S, Colasanti J. Department of Plant and Microbial Biology, University of California, Berkeley, and the Plant Gene Expression Center, Albany, CA 94720, USA. The INDETERMINATE protein, ID1, plays a key role in regulating the transition to flowering in maize. ID1 is the founding member of a plant-specific zinc finger protein family that is defined by a highly conserved amino sequence called the ID domain. The ID domain includes a cluster of three different types of zinc fingers separated from a fourth C2H2 finger by a long spacer; ID1 is distinct from other ID domain proteins by having a much longer spacer. In vitro DNA selection and amplification binding assays and DNA binding experiments showed that ID1 binds selectively to an 11 bp consensus motif via the ID domain. Unexpectedly, site-directed mutagenesis of the ID1 protein showed that zinc fingers located at each end of the ID domain are not required for binding to the consensus motif despite the fact that one of these zinc fingers is a canonical C2H2 DNA binding domain. In addition, an ID1 in vitro deletion mutant that lacks the extra spacer between zinc fingers binds the same 11 bp motif as normal ID1, suggesting that all ID domain-containing proteins recognize the same DNA target sequence. Our results demonstrate that maize ID1 and ID domain proteins have novel zinc finger configurations with unique DNA binding properties. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 15020707 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10871372 # MOTIF: [JP] MA0124.1 NKX3-1 ---------------------------------------------- Nucleic Acids Res. 2000 Jun 15;28(12):2389-95. DNA-binding sequence of the human prostate-specific homeodomain protein NKX3.1. Steadman DJ, Giuffrida D, Gelmann EP. Department of Oncology, Lombardi Cancer Center, Georgetown University School of Medicine, 3800 Reservoir Road NW, Washington, DC 20007-2197, USA. NKX3.1 is a member of the NK class of homeodomain proteins and is most closely related to DROSOPHILA: NK-3. NKX3.1 has predominantly prostate-specific expression in the adult human. Previous studies suggested that NKX3.1 exerts a growth-suppressive effect on prostatic epithelial cells and controls differentiated glandular functions. Using a binding site selection assay with recombinant NKX3.1 protein we identified a TAAGTA consensus binding sequence that has not been reported for any other NK class homeoprotein. By electromobility shift assay we demonstrated that NKX3.1 preferentially binds the TAAGTA sequence rather than the binding site for Nkx2.1 (CAAGTG) or Msx1 (TAATTG). Using mutated binding sites in competitive gel shift assays, we analyzed the nucleotides in the TAAGTA consensus sequence that are important for NKX3.1 binding. The consensus binding site of a naturally occurring polymorphic NKX3.1 protein with arginine replaced by cysteine at position 52 was identical to the wild-type binding sequence. The binding affinities of wild-type and polymorphic NKX3.1 for the TAAGTA consensus site were very similar, with values of 20 and 22 nM, respectively. Wild-type and polymorphic NKX3.1 specifically repressed transcription of luciferase from a reporter vector with three copies of the NKX3.1-binding site upstream from a thymidine kinase promoter. The data show that among NK family proteins NKX3.1 binds a novel DNA sequence and can behave as an in vitro transcriptional repressor. Publication Types: Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S. PMID: 10871372 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12215502 # MOTIF: [JP] MA0121.1 ARR10 ---------------------------------------------- Plant Cell. 2002 Sep;14(9):2015-29. Molecular structure of the GARP family of plant Myb-related DNA binding motifs of the Arabidopsis response regulators. Hosoda K, Imamura A, Katoh E, Hatta T, Tachiki M, Yamada H, Mizuno T, Yamazaki T. Biochemistry Department, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602, Japan. The B motif is a signature of type-B response regulators (ARRs) involved in His-to-Asp phosphorelay signal transduction systems in Arabidopsis. Homologous motifs occur widely in the GARP family of plant transcription factors. To gain general insight into the structure and function of B motifs (or GARP motifs), we characterized the B motif derived from a representative ARR, ARR10, which led to a number of intriguing findings. First, the B motif of ARR10 (named ARR10-B and extending from Thr-179 to Ser-242) possesses a nuclear localization signal, as indicated by the intracellular localization of a green fluorescent protein-ARR10-B fusion protein in onion epidermal cells. Second, the purified ARR10-B molecule binds specifically in vitro to DNA with the core sequence AGATT. This was demonstrated by several in vitro approaches, including PCR-assisted DNA binding site selection, gel retardation assays, and surface plasmon resonance analysis. Finally, the three-dimensional structure of ARR10-B in solution was determined by NMR spectroscopy, showing that it contains a helix-turn-helix structure. Furthermore, the mode of interaction between ARR10-B and the target DNA was assessed extensively by NMR spectroscopy. Together, these results lead us to propose that the mechanism of DNA recognition by ARR10-B is essentially the same as that of homeodomains. We conclude that the B motif is a multifunctional domain responsible for both nuclear localization and DNA binding and suggest that these insights could be applicable generally to the large GARP family of plant transcription factors. Publication Types: Research Support, Non-U.S. Gov't PMID: 12215502 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8530439 # MOTIF: [JP] POL013.1 MED-1 ---------------------------------------------- J Biol Chem. 1995 Dec 22;270(51):30249-52. A conserved downstream element defines a new class of RNA polymerase II promoters. Ince TA, Scotto KW. Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA. Although many TATA-less promoters transcribed by RNA polymerase II initiate transcription at multiple sites, the regulation of multiple start site utilization is not understood. Beginning with the prediction that multiple start site promoters may share regulatory features and using the P-glycoprotein promoter (which can utilize either a single or multiple transcription start site(s)) as a model, several promoters with analogous transcription windows were grouped and searched for the presence of a common DNA element. A downstream protein-binding sequence, MED-1 (Multiple start site Element Downstream), was found in the majority of promoters analyzed. Mutation of this element within the P-glycoprotein promoter reduced transcription by selectively decreasing utilization of downstream start sites. We propose that a new class of RNA polymerase II promoters, those that can utilize a distinctive window of multiple start sites, is defined by the presence of a downstream MED-1 element. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8530439 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7816599 # MOTIF: [JP] MA0095.1 YY1 ---------------------------------------------- Nucleic Acids Res. 1994 Dec 11;22(24):5151-5. An analysis of genes regulated by the multi-functional transcriptional regulator Yin Yang-1. Shrivastava A, Calame K. Department of Biochemistry and Molecular Biophysics, Columbia University College of Physicians and Surgeons, New York, NY 10032. Publication Types: Research Support, Non-U.S. Gov't Review PMID: 7816599 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7632923 # MOTIF: [JP] MA0001.1 AGL3 ---------------------------------------------- Plant Mol Biol. 1995 Jun;28(3):549-67. The Arabidopsis MADS-box gene AGL3 is widely expressed and encodes a sequence-specific DNA-binding protein. Huang H, Tudor M, Weiss CA, Hu Y, Ma H. Cold Spring Harbor Laboratory, NY 11724, USA. The Arabidopsis AGL3 gene was previously identified on the basis of sequence similarity to the floral homeotic gene AGAMOUS (AG), which encodes a protein with a conserved MADS domain that is also found in human and yeast transcription factors (SRF and MCM1, respectively). Analysis of newly isolated full-length cDNA clones as well as genomic clones indicates that AGL3 is indeed a MADS-box gene with a general intron-exon structure similar to other plant MADS-box genes. However, unlike the others, which are expressed specifically in flowers, AGL3 is expressed in all above-ground vegetative organs, as well as in flowers, but not in roots. Furthermore, since AGL3 is MADS-domain protein, it is likely that it is also a DNA-binding protein regulating transcription. To characterize AGL3 as a DNA-binding protein in vitro, we expressed the AGL3 protein in Escherichia coli, and characterized its DNA-binding properties. We show that AGL3 binds to sequences which resemble the target sequences of SRF and MCM1, and have determined the consensus sequence to which AGL3 binds using random oligonucleotides. These results suggest that AGL3 is a widely distributed DNA-binding protein, which may be involved the transcriptional regulation of genes in many cells. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 7632923 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10069063 # MOTIF: [JP] MA0034.1 Gamyb ---------------------------------------------- Plant J. 1999 Jan;17(1):1-9. Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone. Gubler F, Raventos D, Keys M, Watts R, Mundy J, Jacobsen JV. Co-operative Research Centre for Plant Science, Canberra City, ACT, Australia. f.gubler@pi.csiro.au GAMYB is an MYB transcription factor which is expressed in cereal aleurone cells in response to gibberellin (GA). HvGAMYB binds to the TAACAAA box of a high-pl alpha-amylase gene promoter and transcriptionally activates its expression. In this study, we examined the role of HvGAMYB in activating expression of other GA-regulated genes encoding hydrolytic enzymes. In transient expression experiments, HvGAMYB transactivated expression of reporter genes fused to a low-pl alpha-amylase gene promoter, an EII (1-3, 1-4)-beta-glucanase gene promoter and a cathepsin B-like protease promoter. HvGAMYB DNA binding specificity was determined using a PCR-based random site selection using HvGAMYB fusion protein isolated from E. coli. The deduced consensus closely resembled gibberellin response elements in alpha-amylase promoters. Functional analysis of HvGAMYB by transient expression of C terminal HvGAMYB deletions in barley aleurone cells identified two transcriptional activation domains (TADs) which function in transcriptional regulation of both high- and low-pl alpha-amylase promoters. The same TADs were identified using a heterologous yeast expression system. Together, these results indicate that HvGAMYB has two TADs. These domains are C-terminal to its DNA-binding domain. Publication Types: Research Support, Non-U.S. Gov't PMID: 10069063 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8321207 # MOTIF: [JP] MA0035.1 Gata1 # MOTIF: [JP] MA0036.1 GATA2 # MOTIF: [JP] MA0037.1 GATA3 ---------------------------------------------- Mol Cell Biol. 1993 Jul;13(7):3999-4010. DNA-binding specificity of GATA family transcription factors. Merika M, Orkin SH. Division of Hematology/Oncology, Children's Hospital, Dana-Farber Cancer Institute, Boston, Massachusetts. GATA-binding proteins constitute a family of transcription factors that recognize a target site conforming to the consensus WGATAR (W = A or T and R = A or G). Here we have used the method of polymerase chain reaction-mediated random site selection to assess in an unbiased manner the DNA-binding specificity of GATA proteins. Contrary to our expectations, we show that GATA proteins bind a variety of motifs that deviate from the previously assigned consensus. Many of the nonconsensus sequences bind protein with high affinity, equivalent to that of conventional GATA motifs. By using the selected sequences as probes in the electrophoretic mobility shift assay, we demonstrate overlapping, but distinct, sequence preferences for GATA family members, specified by their respective DNA-binding domains. Furthermore, we provide additional evidence for interaction of amino and carboxy fingers of GATA-1 in defining its binding site. By performing cotransfection experiments, we also show that transactivation parallels DNA binding. A chimeric protein containing the finger domain of areA and the activation domains of GATA-1 is capable of activating transcription in mammalian cells through GATA motifs. Our findings suggest a mechanism by which GATA proteins might selectively regulate gene expression in cells in which they are coexpressed. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8321207 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8190643 # MOTIF: [JP] MA0084.1 SRY ---------------------------------------------- Nucleic Acids Res. 1994 Apr 25;22(8):1500-1. Definition of a consensus DNA binding site for SRY. Harley VR, Lovell-Badge R, Goodfellow PN. Department of Genetics, University of Cambridge, UK. Publication Types: Research Support, Non-U.S. Gov't PMID: 8190643 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9571165 # MOTIF: [JP] MA0117.1 Mafb ---------------------------------------------- Biochem Biophys Res Commun. 1998 Apr 17;245(2):412-8. Rat maf-related factors: the specificities of DNA binding and heterodimer formation. Matsushima-Hibiya Y, Nishi S, Sakai M. Department of Biochemistry, School of Medicine, Hokkaido University, Sapporo, Japan. maf is a family of genes encoding bZIP transcription factors. We isolated two cellular maf-related cDNAs, maf-1 (mafB) and maf-2 (c-maf), from rat and determined the specificities of DNA binding and heterodimer formation. Although both Mafs strongly bind to MARE, the consensus Maf recognition sequence (MARE, -TGCTGACTCAGCA-), originally identified by v-Maf protein Maf-1, recognizes a number of sequences containing only the first half of the MARE, -GCTGAC-. On the other hand, no such consensus short sequence could be determined for Maf-2. We determined the specificities of heterodimer formation with all members of the Jun and Fos family. In contrast to v-Maf which forms heterodimers with all Jun and Fos proteins, Maf-1 heterodimerizes with all four Fos proteins, but not at all with the three Jun proteins. Maf-2 heterodimerizes with c-Fos. We have also found that heterodimer formation of Maf-2 with c-Fos dramatically changes the specificity of DNA binding and trans-activation activity from that of the Maf-2 homodimer. These results show that Maf-1 and Maf-2 have significantly different properties and they might have different target genes and functions, in spite of the similarity of their bZip domain structure. Publication Types: Research Support, Non-U.S. Gov't PMID: 9571165 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12746429 # MOTIF: [JP] MA0122.1 Nkx3-2 ---------------------------------------------- J Biol Chem. 2003 Jul 25;278(30):27532-9. Epub 2003 May 13. Characterization of Nkx3.2 DNA binding specificity and its requirement for somitic chondrogenesis. Kim DW, Kempf H, Chen RE, Lassar AB. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA. We have previously shown that Nkx3.2, a member of the NK class of homeoproteins, functions as a transcriptional repressor to promote somitic chondrogenesis. However, it has not been addressed whether Nkx3.2 can bind to DNA in a sequence-specific manner and whether DNA binding by Nkx3.2 is required for its biological activity. In this work, we employed a DNA binding site selection assay, which identified TAAGTG as a high affinity Nkx3.2 binding sequence. Sequence-specific binding of Nkx3.2 to the TAAGTG motif in vitro was confirmed by electrophoretic mobility shift assays, and mutagenesis of this sequence revealed that HRAGTG (where H represents A, C, or T, and R represents A or G) comprises the consensus DNA binding site for Nkx3.2. Consistent with these findings, the expression of a reporter gene containing reiterated Nkx3.2 binding sites was repressed in vivo by Nkx3.2 co-expression. In addition, we have generated a DNA nonbinding point mutant of Nkx3.2 (Nkx3.2-N200Q), which contains an asparagine to glutamine missense mutation in the homeodomain. Interestingly, despite being defective in DNA binding, Nkx3.2-N200Q still retains its intrinsic transcriptional repressor function. Finally, we demonstrate that unlike wild-type Nkx3.2, Nkx3.2-N200Q is unable to activate the chondrocyte differentiation program in somitic mesoderm, indicating that DNA binding by Nkx3.2 is critical for this factor to induce somitic chondrogenesis. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 12746429 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8114711 # MOTIF: [JP] MA0056.1 MZF1_1-4 # MOTIF: [JP] MA0057.1 MZF1_5-13 ---------------------------------------------- Mol Cell Biol. 1994 Mar;14(3):1786-95. Characterization of the DNA-binding properties of the myeloid zinc finger protein MZF1: two independent DNA-binding domains recognize two DNA consensus sequences with a common G-rich core. Morris JF, Hromas R, Rauscher FJ 3rd. Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104. The myeloid zinc finger gene 1, MZF1, encodes a transcription factor which is expressed in hematopoietic progenitor cells that are committed to myeloid lineage differentiation. MZF1 contains 13 C2H2 zinc fingers arranged in two domains which are separated by a short glycine- and proline-rich sequence. The first domain consists of zinc fingers 1 to 4, and the second domain is formed by zinc fingers 5 to 13. We have determined that both sets of zinc finger domains bind DNA. Purified, recombinant MZF1 proteins containing either the first set of zinc fingers or the second set were prepared and used to affinity select DNA sequences from a library of degenerate oligonucleotides by using successive rounds of gel shift followed by PCR amplification. Surprisingly, both DNA-binding domains of MZF1 selected similar DNA-binding consensus sequences containing a core of four or five guanine residues, reminiscent of an NF-kappa B half-site: 1-4, 5'-AGTGGGGA-3'; 5-13, 5'-CGGGnGAGGGGGAA-3'. The full-length MZF1 protein containing both sets of zinc finger DNA-binding domains recognizes synthetic oligonucleotides containing either the 1-4 or 5-13 consensus binding sites in gel shift assays. Thus, we have identified the core DNA consensus binding sites for each of the two DNA-binding domains of a myeloid-specific zinc finger transcription factor. Identification of these DNA-binding sites will allow us to identify target genes regulated by MZF1 and to assess the role of MZF1 as a transcriptional regulator of hematopoiesis. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8114711 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 2329577 # MOTIF: [JP] MA0108.1 TBP # MOTIF: [JP] MA0108.2 TBP # MOTIF: [JP] POL002.1 INR # MOTIF: [JP] POL003.1 GC-box # MOTIF: [JP] POL004.1 CCAAT-box # MOTIF: [JP] POL012.1 TATA-Box ---------------------------------------------- J Mol Biol. 1990 Apr 20;212(4):563-78. Weight matrix descriptions of four eukaryotic RNA polymerase II promoter elements derived from 502 unrelated promoter sequences. Bucher P. Department of Polymer Research, Weizmann Institute of Science, Rehovot, Israel. Optimized weight matrices defining four major eukaryotic promoter elements, the TATA-box, cap signal, CCAAT-, and GC-box, are presented; they were derived by comparative sequence analysis of 502 unrelated RNA polymerase II promoter regions. The new TATA-box and cap signal descriptions differ in several respects from the only hitherto available base frequency Tables. The CCAAT-box matrix, obtained with no prior assumption but CCAAT being the core of the motif, reflects precisely the sequence specificity of the recently discovered nuclear factor NY-I/CP1 but does not include typical recognition sequences of two other purported CCAAT-binding proteins, CTF and CBP. The GC-box description is longer than the previously proposed consensus sequences but is consistent with Sp1 protein-DNA binding data. The notion of a CACCC element distinct from the GC-box seems not to be justified any longer in view of the new weight matrix. Unlike the two fixed-distance elements, neither the CCAAT- nor the GC-box occurs at significantly high frequency in the upstream regions of non-vertebrate genes. Preliminary attempts to predict promoters with the aid of the new signal descriptions were unexpectedly successful. The new TATA-box matrix locates eukaryotic transcription initiation sites as reliably as do the best currently available methods to map Escherichia coli promoters. This analysis was made possible by the recently established Eukaryotic Promoter Database (EPD) of the EMBL Nucleotide Sequence Data Library. In order to derive the weight matrices, a novel algorithm has been devised that is generally applicable to sequence motifs positionally correlated with a biologically defined position in the sequences. The signal must be sufficiently over-represented in a particular region relative to the given site, but need not be present in all members of the input sequence collection. The algorithm iteratively redefines the set of putative motif representatives from which a weight matrix is derived, so as to maximize a quantitative measure of local over-representation, an optimization criterion that naturally combines structural and positional constancy. A comprehensive description of the technique is presented in Methods and Data. PMID: 2329577 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17540862 # MOTIF: [JP] MA0138.2 REST ---------------------------------------------- Science. 2007 Jun 8;316(5830):1497-502. Epub 2007 May 31. Comment in: Science. 2007 Jun 8;316(5830):1441-2. Genome-wide mapping of in vivo protein-DNA interactions. Johnson DS, Mortazavi A, Myers RM, Wold B. Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305-5120, USA. In vivo protein-DNA interactions connect each transcription factor with its direct targets to form a gene network scaffold. To map these protein-DNA interactions comprehensively across entire mammalian genomes, we developed a large-scale chromatin immunoprecipitation assay (ChIPSeq) based on direct ultrahigh-throughput DNA sequencing. This sequence census method was then used to map in vivo binding of the neuron-restrictive silencer factor (NRSF; also known as REST, for repressor element-1 silencing transcription factor) to 1946 locations in the human genome. The data display sharp resolution of binding position [+/-50 base pairs (bp)], which facilitated our finding motifs and allowed us to identify noncanonical NRSF-binding motifs. These ChIPSeq data also have high sensitivity and specificity [ROC (receiver operator characteristic) area >/= 0.96] and statistical confidence (P <10(-4)), properties that were important for inferring new candidate interactions. These include key transcription factors in the gene network that regulates pancreatic islet cell development. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 17540862 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 2208281 # MOTIF: [JP] MA0026.1 Eip74EF ---------------------------------------------- Cell. 1990 Oct 5;63(1):47-61. Molecular interactions within the ecdysone regulatory hierarchy: DNA binding properties of the Drosophila ecdysone-inducible E74A protein. Urness LD, Thummel CS. Howard Hughes Medical Institute, Department of Human Genetics, University of Utah Medical Center, Salt Lake City 84132. The E74 early ecdysone-inducible gene plays a key role in the regulatory hierarchy activated by ecdysone at the onset of Drosophila metamorphosis. We show here that E74A protein binds to three adjacent sites in the middle of the E74 gene. The consensus sequence for E74A protein binding, determined by random-sequence oligonucleotide selection, contains an invariant purine-rich core sequence, C/AGGAA. This sequence is also present in the binding sites of two mammalian proteins that, like E74A, are related to the ets oncoprotein. Antibody staining of larval salivary gland polytene chromosomes revealed that E74A protein binds to both early and late ecdysone-inducible puffs. This study supports Ashburner's proposal that the early puffs encode site-specific DNA binding proteins that directly interact with the early and late ecdysone-inducible puffs. Publication Types: Research Support, Non-U.S. Gov't PMID: 2208281 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1406630 # MOTIF: [JP] MA0101.1 REL # MOTIF: [JP] MA0105.1 NFKB1 # MOTIF: [JP] MA0107.1 RELA ---------------------------------------------- Mol Cell Biol. 1992 Oct;12(10):4412-21. Selection of optimal kappa B/Rel DNA-binding motifs: interaction of both subunits of NF-kappa B with DNA is required for transcriptional activation. Kunsch C, Ruben SM, Rosen CA. Department of Gene Regulation, Roche Institute of Molecular Biology, Nutley, New Jersey 07110. Analysis of the p50 and p65 subunits of the NF-kappa B transcription factor complex has revealed that both proteins can interact with related DNA sequences through either homo- or heterodimer formation. In addition, the product of the proto-oncogene c-rel can bind to similar DNA motifs by itself or as a heterodimer with p50 or p65. However, these studies have used a limited number of known kappa B DNA motifs, and the question of the optimal DNA sequences preferred by each homodimer has not been addressed. Using purified recombinant p50, p65, and c-Rel proteins, optimal DNA-binding motifs were selected from a pool of random oligonucleotides. Alignment of the selected sequences allowed us to predict a consensus sequence for binding of the individual homodimeric Rel-related proteins, and DNA-protein binding analysis of the selected DNA sequences revealed sequence specificity of the proteins. Contrary to previous assumptions, we observed that p65 homodimers can interact with a subset of DNA sequences not recognized by p50 homodimers. Differential binding affinities were also obtained with p50- and c-Rel-selected sequences. Using either a p50- or p65-selected kappa B motif, which displayed differential binding with respect to the other protein, little to no binding was observed with the heterodimeric NF-kappa B complex. Similarly, in transfection experiments in which the selective kappa B binding sites were used to drive the expression of a chloramphenicol acetyltransferase reporter construct, the p65- and p50-selected motifs were activated only in the presence of p65 and p50/65 (a chimeric protein with the p50 DNA binding domain and p65 activation domain) expression vectors, respectively, and neither demonstrated a significant response to stimuli that induce NF-kappa B activity. These findings demonstrate that interaction of both subunits of the heterodimeric NF-kappa B complex with DNA is required for DNA binding and transcriptional activation and suggest that transcriptional activation mediated by the individual rel-related proteins will differ dramatically, depending on the specific kappa B motifs present. Publication Types: Research Support, Non-U.S. Gov't PMID: 1406630 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16234508 # MOTIF: [JP] MA0259.1 HIF1A::ARNT ---------------------------------------------- Sci STKE. 2005 Oct 18;2005(306):re12. Integration of oxygen signaling at the consensus HRE. Wenger RH, Stiehl DP, Camenisch G. Institute of Physiology, Center for Integrative Human Physiology, University of Zurich, CH-8057 Zurich, Switzerland. roland.wenger@access.unizh.ch The hypoxia-inducible factor 1 (HIF-1) was initially identified as a transcription factor that regulated erythropoietin gene expression in response to a decrease in oxygen availability in kidney tissue. Subsequently, a family of oxygen-dependent protein hydroxylases was found to regulate the abundance and activity of three oxygen-sensitive HIFalpha subunits, which, as part of the HIF heterodimer, regulated the transcription of at least 70 different effector genes. In addition to responding to a decrease in tissue oxygenation, HIF is proactively induced, even under normoxic conditions, in response to stimuli that lead to cell growth, ultimately leading to higher oxygen consumption. The growing cell thus profits from an anticipatory increase in HIF-dependent target gene expression. Growth stimuli-activated signaling pathways that influence the abundance and activity of HIFs include pathways in which kinases are activated and pathways in which reactive oxygen species are liberated. These pathways signal to the HIF protein hydroxylases, as well as to HIF itself, by means of covalent or redox modifications and protein-protein interactions. The final point of integration of all of these pathways is the hypoxia-response element (HRE) of effector genes. Here, we provide comprehensive compilations of the known growth stimuli that promote increases in HIF abundance, of protein-protein interactions involving HIF, and of the known HIF effector genes. The consensus HRE derived from a comparison of the HREs of these HIF effectors will be useful for identification of novel HIF target genes, design of oxygen-regulated gene therapy, and prediction of effects of future drugs targeting the HIF system. Publication Types: Review PMID: 16234508 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16227614 # MOTIF: [JP] POL008.1 DCE_S_I # MOTIF: [JP] POL009.1 DCE_S_II # MOTIF: [JP] POL010.1 DCE_S_III ---------------------------------------------- Mol Cell Biol. 2005 Nov;25(21):9674-86. Erratum in: Mol Cell Biol. 2005 Dec;25(24):11192. Functional characterization of core promoter elements: the downstream core element is recognized by TAF1. Lee DH, Gershenzon N, Gupta M, Ioshikhes IP, Reinberg D, Lewis BA. Department of Biochemistry, Robert Woods Johnson Medical School, 683 Hoes Lane, Piscataway, NJ 08854, USA. Downstream elements are a newly appreciated class of core promoter elements of RNA polymerase II-transcribed genes. The downstream core element (DCE) was discovered in the human beta-globin promoter, and its sequence composition is distinct from that of the downstream promoter element (DPE). We show here that the DCE is a bona fide core promoter element present in a large number of promoters and with high incidence in promoters containing a TATA motif. Database analysis indicates that the DCE is found in diverse promoters, supporting its functional relevance in a variety of promoter contexts. The DCE consists of three subelements, and DCE function is recapitulated in a TFIID-dependent manner. Subelement 3 can function independently of the other two and shows a TFIID requirement as well. UV photo-cross-linking results demonstrate that TAF1/TAF(II)250 interacts with the DCE subelement DNA in a sequence-dependent manner. These data show that downstream elements consist of at least two types, those of the DPE class and those of the DCE class; they function via different DNA sequences and interact with different transcription activation factors. Finally, these data argue that TFIID is, in fact, a core promoter recognition complex. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 16227614 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7687740 # MOTIF: [JP] MA0050.1 IRF1 # MOTIF: [JP] MA0051.1 IRF2 ---------------------------------------------- Mol Cell Biol. 1993 Aug;13(8):4531-8. Recognition DNA sequences of interferon regulatory factor 1 (IRF-1) and IRF-2, regulators of cell growth and the interferon system. Tanaka N, Kawakami T, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. Interferon (IFN) regulatory factor 1 (IRF-1) and IRF-2 were originally identified as transcription factors involved in the regulation of the IFN system. IRF-1 functions as a transcriptional activator, while IRF-2 represses IRF-1 function. More recently, evidence has been provided that IRF-1 and IRF-2 manifest antioncogenic and oncogenic properties, respectively, and that loss of one or both of the IRF-1 alleles may be critical for the development of human hematopoietic neoplasms. Both factors show a high degree of structural similarity in their N-terminal DNA-binding domains, and previous studies suggested that IRF-1 and IRF-2 bind to similar or identical cis elements within type I IFN (IFN-alpha and -beta) and IFN-inducible genes. However, the exact recognition sequences of these two factors have not yet been determined; hence, the spectrum of the IRF-responsive genes remains unclear. In this study, we determined the DNA sequences recognized by IRF-1 and IRF-2, using a polymerase chain reaction-assisted DNA-binding site selection method. We report that sequences selected by this method and the affinities for each sequence were virtually indistinguishable between IRF-1 and IRF-2. We confirm the presence of two contiguous IRF recognition sequences within the promoter region of the IFN-beta gene and of at least one such sequence in all of the IFN-inducible genes examined. Furthermore, we report the presence of potential IRF sequences in the upstream region of several genes involved in cell growth control. Publication Types: In Vitro Research Support, Non-U.S. Gov't PMID: 7687740 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18555785 # MOTIF: [JP] MA0039.2 Klf4 # MOTIF: [JP] MA0104.2 Mycn # MOTIF: [JP] MA0141.1 Esrrb # MOTIF: [JP] MA0142.1 Pou5f1 # MOTIF: [JP] MA0143.1 Sox2 # MOTIF: [JP] MA0144.1 Stat3 # MOTIF: [JP] MA0145.1 Tcfcp2l1 # MOTIF: [JP] MA0146.1 Zfx # MOTIF: [JP] MA0147.1 Myc ---------------------------------------------- Cell. 2008 Jun 13;133(6):1106-17. Integration of external signaling pathways with the core transcriptional network in embryonic stem cells. Chen X, Xu H, Yuan P, Fang F, Huss M, Vega VB, Wong E, Orlov YL, Zhang W, Jiang J, Loh YH, Yeo HC, Yeo ZX, Narang V, Govindarajan KR, Leong B, Shahab A, Ruan Y, Bourque G, Sung WK, Clarke ND, Wei CL, Ng HH. Gene Regulation Laboratory, Genome Institute of Singapore, Singapore 138672. Transcription factors (TFs) and their specific interactions with targets are crucial for specifying gene-expression programs. To gain insights into the transcriptional regulatory networks in embryonic stem (ES) cells, we use chromatin immunoprecipitation coupled with ultra-high-throughput DNA sequencing (ChIP-seq) to map the locations of 13 sequence-specific TFs (Nanog, Oct4, STAT3, Smad1, Sox2, Zfx, c-Myc, n-Myc, Klf4, Esrrb, Tcfcp2l1, E2f1, and CTCF) and 2 transcription regulators (p300 and Suz12). These factors are known to play different roles in ES-cell biology as components of the LIF and BMP signaling pathways, self-renewal regulators, and key reprogramming factors. Our study provides insights into the integration of the signaling pathways into the ES-cell-specific transcription circuitries. Intriguingly, we find specific genomic regions extensively targeted by different TFs. Collectively, the comprehensive mapping of TF-binding sites identifies important features of the transcriptional regulatory networks that define ES-cell identity. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 18555785 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15475254 # MOTIF: [JP] SA0001.1 at_AC_acceptor # MOTIF: [JP] SA0002.1 at_AC_acceptor # MOTIF: [JP] SA0003.1 at_AC_acceptor # MOTIF: [JP] SD0001.1 at_AC_acceptor # MOTIF: [JP] SD0002.1 at_AC_acceptor # MOTIF: [JP] SD0003.1 at_AC_acceptor ---------------------------------------------- Genomics. 2004 Oct;84(4):762-6. Information for the Coordinates of Exons (ICE): a human splice sites database. Chong A, Zhang G, Bajic VB. Institute for Infocomm Research, 21 Heng Mui Keng Terrace, Singapore 119613, Singapore. achong@i2r.a-star.edu.sg We present a comprehensive database, Information for the Coordinates of Exons (ICE), of genomic splice sites (SSs) for 10,803 human genes. ICE contains 91,846 pairs of donor acceptor sites, supported by the alignment of "full-length" human mRNAs (including transcript variants) on human genomic sequences. ICE represents the largest collection of human SSs known to date and provides a significant resource to both molecular biologists and bioinformaticians alike. A user can visualize and extract genomic sequences around SSs of the donor acceptor pairs and can also visualize the primary structure of individual genes. We list in this article the 22 most frequently found canonical and noncanonical splice sites. The top four most represented donor acceptor pairs (GT-AG, GC-AG, AT-AC, and GT-GG) accounted for 99.16% of our data set. In addition, we calculated the SS matrix models for the three most common donor acceptor pairs. The database is focused on providing SSs and surrounding sequence information, associated SS and sequence characteristics, and relation to overall transcript structure. It allows targeted search and presents evidence for the gene structure. Copyright 2004 Elsevier Inc. PMID: 15475254 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7797561 # MOTIF: [JP] MA0063.1 Nkx2-5 ---------------------------------------------- J Biol Chem. 1995 Jun 30;270(26):15628-33. Identification of novel DNA binding targets and regulatory domains of a murine tinman homeodomain factor, nkx-2.5. Chen CY, Schwartz RJ. Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030, USA. A murine cardiac-specific homeodomain gene named csx (Komuro, I., and Izumo. S. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 8145-8149) and nkx-2.5 (Lints, T. J., Parsons, L. M., Hartley, L., Lyons, I., and Harvey, R. P. (1993) Development 119, 419-431) was identified as a potential vertebrate homologue of Drosophila tinman, a mesoderm determination factor required for insect heart formation (Bodmer, R. (1993) Development 118, 719-729). Bacterial expression of the nkx-2.5 homeodomain allowed us to identify downstream DNA targets from a library of randomly generated oligonucleotides. High affinity nkx-2.5 DNA binding sites, 5'-TNNAGTG-3', represented novel binding sequences, whereas intermediate and weaker affinity sites, 5'-C(A/T)TTAATTN-3', contained the typical 5'-TAAT-3' core required by most homeodomain factors for DNA binding. We also observed that nkx-2.5 served as a modest transcription activator in transfection assays done in 10T1/2 fibroblasts with multimerized binding sites linked to a luciferase reporter gene. Functional dissection of nkx-2.5 revealed a COOH-terminal inhibitory domain composed mainly of clusters of alanines and prolines, which appeared to mask a potent activation domain composed of hydrophobic and highly charged amino acids. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 7797561 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8413232 # MOTIF: [JP] MA0002.1 RUNX1 ---------------------------------------------- Mol Cell Biol. 1993 Oct;13(10):6336-45. Identification of AML-1 and the (8;21) translocation protein (AML-1/ETO) as sequence-specific DNA-binding proteins: the runt homology domain is required for DNA binding and protein-protein interactions. Meyers S, Downing JR, Hiebert SW. Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105. The AML1 gene on chromosome 21 is disrupted in the (8;21)(q22;q22) translocation associated with acute myelogenous leukemia and encodes a protein with a central 118-amino-acid domain with 69% homology to the Drosophila pair-rule gene, runt. We demonstrate that AML-1 is a DNA-binding protein which specifically interacts with a sequence belonging to the group of enhancer core motifs, TGT/cGGT. Electrophoretic mobility shift analysis of cell extracts identified two AML-1-containing protein-DNA complexes whose electrophoretic mobilities were slower than those of complexes formed with AML-1 produced in vitro. Mixing of in vitro-produced AML-1 with cell extracts prior to gel mobility shift analysis resulted in the formation of higher-order complexes. Deletion mutagenesis of AML-1 revealed that the runt homology domain mediates both sequence-specific DNA binding and protein-protein interactions. The hybrid product, AML-1/ETO, which results from the (8;21) translocation and retains the runt homology domain, both recognizes the AML-1 consensus sequence and interacts with other cellular proteins. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8413232 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18798982 # MOTIF: [JP] MA0148.1 FOXA1 ---------------------------------------------- Genome Biol. 2008;9(9):R137. Epub 2008 Sep 17. Model-based analysis of ChIP-Seq (MACS). Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nussbaum C, Myers RM, Brown M, Li W, Liu XS. Department of Biostatistics and Computational Biology, Dana-Farber Cancer Institute and Harvard School of Public Health, Boston, MA 02115, USA. We present Model-based Analysis of ChIP-Seq data, MACS, which analyzes data generated by short read sequencers such as Solexa's Genome Analyzer. MACS empirically models the shift size of ChIP-Seq tags, and uses it to improve the spatial resolution of predicted binding sites. MACS also uses a dynamic Poisson distribution to effectively capture local biases in the genome, allowing for more robust predictions. MACS compares favorably to existing ChIP-Seq peak-finding algorithms, and is freely available. Publication Types: Comparative Study Research Support, N.I.H., Extramural PMID: 18798982 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 14752047 # MOTIF: [JP] MA0131.1 MIZF ---------------------------------------------- Nucleic Acids Res. 2004 Jan 29;32(2):590-7. Print 2004. Sequence-specific transcriptional repression by an MBD2-interacting zinc finger protein MIZF. Sekimata M, Homma Y. Department of Biomolecular Sciences, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295, Japan. matam@fmu.ac.jp MBD2 is a member of the methyl-CpG-binding protein family that plays an important role in methylated DNA silencing. We have recently identified a novel zinc finger protein, MIZF, as an MBD2-binding partner. To understand the physiological function of MIZF in MBD2-mediated gene silencing, we investigated the DNA-binding properties of MIZF and its potential target genes. Using a cyclic amplification and selection of targets technique, the consensus sequence CGGACGTT, which contains a conserved CGGAC core, was determined as sufficient for MIZF binding. Deletion of individual zinc fingers revealed that five of the seven zinc fingers are required for DNA binding. Reporter assays demonstrated that MIZF represses transcription from the promoter including this DNA sequence. A database search indicated that a variety of human genes, including Rb, contain this sequence in their promoter region. MIZF actually bound to its recognition sequence within the Rb promoter and repressed Rb transcription. These results suggest that MIZF, through its DNA-binding activity, acts as a sequence-specific transcriptional repressor likely involved in MBD2-mediated epigenetic gene silencing. Publication Types: Research Support, Non-U.S. Gov't PMID: 14752047 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9161031 # MOTIF: [JP] MA0044.1 HMG-1 # MOTIF: [JP] MA0045.1 HMG-I/Y ---------------------------------------------- Plant J. 1997 Apr;11(4):703-15. High mobility group proteins HMG-1 and HMG-I/Y bind to a positive regulatory region of the pea plastocyanin gene promoter. Webster CI, Packman LC, Pwee KH, Gray JC. Cambridge Centre for Molecular Recognition, University of Cambridge, UK. A 268 bp region (P268) of the pea plastocyanin gene promoter responsible for high-level expression has been shown to interact with the high mobility group proteins HMG-1 and HMG-I/Y isolated from pea shoot chromatin. cDNAs encoding an HMG-1 protein of 154 amino acid residues containing a single HMG-box and a C-terminal acidic tail and an HMG-I/Y-like protein of 197 amino acid residues containing four AT-hooks have been isolated and expressed in Escherichia coli to provide large amounts of full-length proteins. DNase I footprinting identified eight binding sites for HMG-I/Y and six binding sites for HMG-1 in P268. Inhibition of binding by the antibiotic distamycin, which binds in the minor groove of A/T-rich DNA, revealed that HMG-I/Y binding was 400-fold more sensitive than HMG-1 binding. Binding-site selection from a pool of random oligonucleotides indicated that HMG-I/Y binds to oligonucleotides containing stretches of five or more A/T bp and HMG-1 binds preferentially to oligonucleotides enriched in dinucleotides such as TpT and TpG. Publication Types: Research Support, Non-U.S. Gov't PMID: 9161031 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 14704343 # MOTIF: [JP] MA0132.1 Pdx1 ---------------------------------------------- Nucleic Acids Res. 2004 Jan 2;32(1):54-64. Print 2004. Role of intrinsic DNA binding specificity in defining target genes of the mammalian transcription factor PDX1. Liberzon A, Ridner G, Walker MD. Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel. PDX1 is a homeodomain transcription factor essential for pancreatic development and mature beta cell function. Homeodomain proteins typically recognize short TAAT DNA motifs in vitro: this binding displays paradoxically low specificity and affinity, given the extremely high specificity of action of these proteins in vivo. To better understand how PDX1 selects target genes in vivo, we have examined the interaction of PDX1 with natural and artificial binding sites. Comparison of PDX1 binding sites in several target promoters revealed an evolutionarily conserved pattern of nucleotides flanking the TAAT core. Using competitive in vitro DNA binding assays, we defined three groups of binding sites displaying high, intermediate and low affinity. Transfection experiments revealed a striking correlation between the ability of each sequence to activate transcription in cultured beta cells, and its ability to bind PDX1 in vitro. Site selection from a pool of oligonucleotides (sequence NNNTAATNNN) revealed a non-random preference for particular nucleotides at the flanking locations, resembling natural PDX1 binding sites. Taken together, the data indicate that the intrinsic DNA binding specificity of PDX1, in particular the bases adjacent to TAAT, plays an important role in determining the spectrum of target genes. Publication Types: Research Support, Non-U.S. Gov't PMID: 14704343 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8096059 # MOTIF: [JP] MA0027.1 En1 ---------------------------------------------- Mol Cell Biol. 1993 Apr;13(4):2354-65. Nucleotides flanking a conserved TAAT core dictate the DNA binding specificity of three murine homeodomain proteins. Catron KM, Iler N, Abate C. Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, New Jersey. Murine homeobox genes play a fundamental role in directing embryogenesis by controlling gene expression during development. The homeobox encodes a DNA binding domain (the homeodomain) which presumably mediates interactions of homeodomain proteins with specific DNA sites in the control regions of target genes. However, the bases for these selective DNA-protein interactions are not well defined. In this report, we have characterized the DNA binding specificities of three murine homeodomain proteins, Hox 7.1, Hox 1.5, and En-1. We have identified optimal DNA binding sites for each of these proteins by using a random oligonucleotide selection strategy. Comparison of the sequences of the selected binding sites predicted a common consensus site that contained the motif (C/G)TAATTG. The TAAT core was essential for DNA binding activity, and the nucleotides flanking this core directed binding specificity. Whereas variations in the nucleotides flanking the 5' side of the TAAT core produced modest alterations in binding activity for all three proteins, perturbations of the nucleotides directly 3' of the core distinguished the binding specificity of Hox 1.5 from those of Hox 7.1 and En-1. These differences in binding activity reflected differences in the dissociation rates rather than the equilibrium constants of the protein-DNA complexes. Differences in DNA binding specificities observed in vitro may contribute to selective interactions of homeodomain proteins with potential binding sites in the control regions of target genes. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8096059 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15563547 # MOTIF: [JP] MA0112.1 ESR1 # MOTIF: [JP] MA0113.1 NR3C1 ---------------------------------------------- Mol Endocrinol. 2005 Mar;19(3):595-606. Epub 2004 Nov 24. Prediction of nuclear hormone receptor response elements. Sandelin A, Wasserman WW. Center for Genomics and Bioinformatics, Karolinska Institutet, Stockholm, Sweden. The nuclear receptor (NR) class of transcription factors controls critical regulatory events in key developmental processes, homeostasis maintenance, and medically important diseases and conditions. Identification of the members of a regulon controlled by a NR could provide an accelerated understanding of development and disease. New bioinformatics methods for the analysis of regulatory sequences are required to address the complex properties associated with known regulatory elements targeted by the receptors because the standard methods for binding site prediction fail to reflect the diverse target site configurations. We have constructed a flexible Hidden Markov Model framework capable of predicting NHR binding sites. The model allows for variable spacing and orientation of half-sites. In a genome-scale analysis enabled by the model, we show that NRs in Fugu rubripes have a significant cross-regulatory potential. The model is implemented in a web interface, freely available for academic researchers, available at http://mordor.cgb.ki.se/NHR-scan. Publication Types: Research Support, Non-U.S. Gov't PMID: 15563547 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15634773 # MOTIF: [JP] MA0164.1 Nr2e3 ---------------------------------------------- J Neurosci. 2005 Jan 5;25(1):118-29. The rod photoreceptor-specific nuclear receptor Nr2e3 represses transcription of multiple cone-specific genes. Chen J, Rattner A, Nathans J. Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. This study addresses one genetic regulatory mechanism that establishes the distinct identities of rod and cone photoreceptors. Previous work has shown that mutations in either humans or mice in the gene coding for photoreceptor-specific nuclear receptor Nr2e3 cause a progressive retinal degeneration characterized by increased numbers of short-wave cones. In the present work, we have examined the cellular and developmental pattern of Nr2e3 protein localization in mammals and fish, identified an optimal Nr2e3 DNA-binding site using cycles of binding to recombinant Nr2e3, characterized the transcriptional activity of wild type and one of the disease-associated point mutations in Nr2e3 in transfected cells, and characterized the transcriptional defects in the naturally occurring Nr2e3 mutant (rd7) mouse. These experiments indicate that in the mature vertebrate retina Nr2e3 is expressed exclusively in rods, that expression of Nr2e3 is one of the earliest events in the pathway of rod-specific photoreceptor development, and that Nr2e3 functions, either directly or indirectly, as a repressor of cone-specific genes in rod photoreceptor cells. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 15634773 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 11165476 # MOTIF: [JP] MA0111.1 Spz1 ---------------------------------------------- Mech Dev. 2001 Feb;100(2):177-87. Spz1, a novel bHLH-Zip protein, is specifically expressed in testis. Hsu SH, Shyu HW, Hsieh-Li HM, Li H. Institute of Molecular Biology, Academia Sinica, 11529, Taipei, Taiwan. We isolated a novel bHLH-Zip gene designated Spz1 from a mouse testis cDNA library. Spz1 is expressed specifically in the testis and epididymis. Immunofluorescence staining detected Spz1 protein in the nuclei of LFG6 Leydig cells. The ability of Spz1 protein to bind to the bHLH consensus-binding site, the E-box, was confirmed by EMSA, and a 9-bp asymmetric target site was identified by random selection and PCR amplification. Hormonal regulation of Spz1 was investigated and downregulation of Spz1 expression by testosterone and retinoic acid was found. This nuclear transcription factor may play a crucial role in spermatogenesis by regulating cell proliferation or differentiation through binding to specific DNA sequences like other bHLH-Zip molecules. Publication Types: Research Support, Non-U.S. Gov't PMID: 11165476 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7901837 # MOTIF: [JP] MA0075.1 Prrx2 ---------------------------------------------- Nucleic Acids Res. 1993 Oct 11;21(20):4711-20. DNA-binding specificity of the S8 homeodomain. de Jong R, van der Heijden J, Meijlink F. Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Utrecht. The murine S8 homeobox gene is expressed in a mesenchyme-specific pattern in embryos, as well as in mesodermal cell lines. The S8 homeodomain is overall similar to paired type homeodomains, but at position 50, which is crucial for specific DNA recognition, it contains a Gln, as is found in Antennapedia (Antp)-type homeodomains. We determined the DNA-binding specificity of the purified S8 homeodomain by in vitro selection of random oligonucleotides. The resulting 11-bp consensus binding site, ANC/TC/TAATTAA/GC resembles, but subtly differs from, the recognition sequences of Antp-type homeodomains. Equilibrium binding constants of down to 6.0 x 10(-10) M were found for binding of the S8 homeodomain to selected oligonucleotides. Using specific antibodies and an oligonucleotide containing an S8-site, we detected by band-shift two abundant DNA binding activities in mesodermal cell lines that correspond to S8 and two more that correspond to its close relative MHox. These S8 protein forms are differentially expressed in retinoic acid-treated P19 EC cells. PMID: 7901837 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7901838 # MOTIF: [JP] MA0005.1 AG ---------------------------------------------- Nucleic Acids Res. 1993 Oct 11;21(20):4769-76. Isolation and characterization of the binding sequences for the product of the Arabidopsis floral homeotic gene AGAMOUS. Huang H, Mizukami Y, Hu Y, Ma H. Cold Spring Harbor Laboratory, NY 11724-2212. The Arabidopsis floral homeotic gene AGAMOUS (AG) is required for normal flower development. The deduced AG protein contains a region which shares substantial sequence similarity with the DNA-binding domains of known transcription factors, SRF (human) and MCM1 (yeast). Therefore, it is likely that AG is also a DNA-binding protein regulating transcription of floral genes. We describe here several experiments to characterize AG-DNA binding in vitro. We show that AG indeed binds a DNA sequence matching the consensus of SRF targets. Further, we have selected the AG-binding sequences from a pool of random oligonucleotides, and deduced an AG-binding consensus sequence of TT(A/T)CC(A/T)(A/t)2(T/A)NNGG(-G)(A/t)2. We have demonstrated that AG binds to the consensus region of three of the oligonucleotides by footprinting analysis. Finally, we have examined AG's relative binding affinity for different sequences, as compared to SRF, by gel mobility shift analysis. Our results indicate that AG is a sequence-specific DNA-binding protein, and that the AG-binding consensus sequence is similar to those of MCM1 and SRF. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 7901838 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1425594 # MOTIF: [JP] MA0028.1 ELK1 ---------------------------------------------- EMBO J. 1992 Dec;11(12):4631-40. Spatial flexibility in ternary complexes between SRF and its accessory proteins. Treisman R, Marais R, Wynne J. Transcription Laboratory, Imperial Cancer Research Fund, Lincoln's Inn Fields, London, UK. We investigated the sequence requirements for ternary complex formation by the transcription factor SRF and its Ets domain accessory factors Elk-1 and SAP-1. Ternary complex formation is specified by an SRF consensus site CC(A/T)6GG and a neighbouring Ets motif (C/A)(C/A)GGA(A/T), which is contacted by Elk-1/SAP-1. Both the spacing of these sequences and their relative orientation can be substantially altered with little effect on the efficiency of ternary complex formation. Efficient ternary complex formation by Elk-1 is mediated by the B box, a conserved 21 amino acid region located 50 residues C-terminal to the Ets domain, which also acts to inhibit autonomous DNA binding. Binding studies with the isolated Ets domains indicate that ternary complex formation compensates for low affinity Ets domain-DNA interactions. Several naturally occurring SREs containing Ets motifs at different locations to that in the human c-fos SRE allow SAP-1 and Elk-1 recruitment in vitro. We discuss the mechanism of ternary complex formation. PMID: 1425594 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 11139380 # MOTIF: [JP] MA0065.1 PPARG::RXRA # MOTIF: [JP] MA0066.1 PPARG ---------------------------------------------- Biochem J. 2001 Jan 15;353(Pt 2):193-8. Dual DNA-binding specificity of peroxisome-proliferator-activated receptor gamma controlled by heterodimer formation with retinoid X receptor alpha. Okuno M, Arimoto E, Ikenobu Y, Nishihara T, Imagawa M. Laboratory of Environmental Biochemistry, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamada-Oka, Suita, Osaka 565-0871, Japan. The peroxisome-proliferator-activated receptor gamma (PPARgamma) is a member of the steroid/thyroid nuclear receptor superfamily of ligand-activated transcription factors. PPARgamma forms a heterodimer with the retinoid X receptor alpha (RXRalpha) and binds to a common consensus response element consisting of a direct repeat of two hexanucleotides spaced by one nucleotide (DR1 motif). However, other hexamer configurations for binding of PPARgamma have not been considered. By using PCR-mediated random site selection, the DNA sequence preferences for PPARgamma binding were examined. In this study, we have demonstrated that PPARgamma has dual DNA-binding specificity; binding to both the DR1 motif and a palindromic sequence with three bases as spacers (Pal3 motif). The consensus sequence selected by equimolar amounts of PPARgamma and RXRalpha was a perfect DR1 motif, whereas a relatively large population of Pal3 was observed when a 30-fold molar excess of PPARgamma over RXRalpha was used. Gel-shift analysis revealed that the PPARgamma homodimer could bind to Pal3 and that the affinity constant of the PPARgamma homodimer for Pal3 was nearly the same as that of the PPARgamma/RXRalpha heterodimer for DR1. The addition of RXRalpha decreased the binding affinity of PPARgamma for Pal3, indicating that the DNA-binding specificity of PPARgamma could be altered by heterodimer formation with RXRalpha. Publication Types: Research Support, Non-U.S. Gov't PMID: 11139380 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1396566 # MOTIF: [JP] MA0087.1 Sox5 ---------------------------------------------- EMBO J. 1992 Oct;11(10):3705-12. An SRY-related gene expressed during spermatogenesis in the mouse encodes a sequence-specific DNA-binding protein. Denny P, Swift S, Connor F, Ashworth A. Chester Beatty Laboratories, Institute of Cancer Research, London, UK. SRY, the testis determining gene, encodes a member of a family of DNA binding proteins characterized by an amino acid sequence motif known as the HMG box. Using degenerate primers and the polymerase chain reaction, we have isolated SRY-related cDNAs from adult murine testis RNA. One of these, Sox-5, encodes a 43 kDa HMG-box protein with similarities to transcription activating proteins. Anti-Sox-5 antibody was used to analyse expression of Sox-5 in pre-pubertal testis and in fractionated spermatogenic cells. Sox-5 is restricted to post-meiotic germ cells, being found at highest levels in round spermatids. Sox-5 is a DNA binding protein and binding site selection assays suggest that it can bind specifically to oligonucleotides containing the consensus motif AACAAT. Sry can also bind to this motif, indicating that the Sry family may have overlapping sequence specificities. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 1396566 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8289805 # MOTIF: [JP] MA0091.1 TAL1::TCF3 ---------------------------------------------- Mol Cell Biol. 1994 Feb;14(2):1256-65. Preferred sequences for DNA recognition by the TAL1 helix-loop-helix proteins. Hsu HL, Huang L, Tsan JT, Funk W, Wright WE, Hu JS, Kingston RE, Baer R. Department of Microbiology, University of Texas Southwestern Medical Center, Dallas 75235. Tumor-specific activation of the TAL1 gene is the most common genetic alteration seen in patients with T-cell acute lymphoblastic leukemia. The TAL1 gene products contain the basic helix-loop-helix (bHLH) domain, a protein dimerization and DNA-binding motif common to several known transcription factors. A binding-site selection procedure has now been used to evaluate the DNA recognition properties of TAL1. These studies demonstrate that TAL1 polypeptides do not have intrinsic DNA-binding activity, presumably because of their inability to form bHLH homodimers. However, TAL1 readily interacts with any of the known class A bHLH proteins (E12, E47, E2-2, and HEB) to form heterodimers that bind DNA in a sequence-specific manner. The TAL1 heterodimers preferentially recognize a subset of E-box elements (CANNTG) that can be represented by the consensus sequence AACAGATGGT. This consensus is composed of half-sites for recognition by the participating class A bHLH polypeptide (AACAG) and the TAL1 polypeptide (ATGGT). TAL1 heterodimers with DNA-binding activity are readily detected in nuclear extracts of Jurkat, a leukemic cell line derived from a patient with T-cell acute lymphoblastic leukemia. Hence, TAL1 is likely to bind and regulate the transcription of a unique subset of subordinate target genes, some of which may mediate the malignant function of TAL1 during T-cell leukemogenesis. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8289805 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8289804 # MOTIF: [JP] MA0048.1 NHLH1 ---------------------------------------------- Mol Cell Biol. 1994 Feb;14(2):1245-55. HEN1 encodes a 20-kilodalton phosphoprotein that binds an extended E-box motif as a homodimer. Brown L, Baer R. Department of Microbiology, University of Texas Southwestern Medical Center, Dallas 75235. HEN1 and HEN2 encode neuron-specific polypeptides that contain the basic helix-loop-helix (bHLH) motif, a protein dimerization and DNA-binding domain common to several known transcription factors. We now describe characteristics of the HEN1 gene product that are consistent with its postulated role as a transcription factor that functions during development of the mammalian nervous system. Thus, transcription of the HEN1 gene is activated upon the induction of neural differentiation in PC12 cells by nerve growth factor. HEN1 encodes a 20-kDa polypeptide (pp20HEN1) that is phosphorylated exclusively at serine residues and forms dimeric bHLH complexes either by self-association or by heterologous interaction with the E2A gene products (E12 or E47). The resultant HEN1/HEN1 homodimers and HEN1/E2A heterodimers bind DNA in a sequence-specific manner. Moreover, a binding site selection procedure revealed that HEN1-HEN1 homodimers preferentially recognize E-box motifs represented by an 18-bp consensus sequence (GGGNCG CAGCTGCGNCCC). The E-box half-site recognized by HEN1 polypeptides (GGGNCGCAG) is distinct from those of other known bHLH proteins, suggesting that HEN1 binds, an regulates the transcription of, a unique subset of target genes during neural development. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8289804 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8657121 # MOTIF: [JP] MA0019.1 Ddit3::Cebpa ---------------------------------------------- Mol Cell Biol. 1996 Apr;16(4):1479-89. Stress-induced binding of the transcriptional factor CHOP to a novel DNA control element. Ubeda M, Wang XZ, Zinszner H, Wu I, Habener JF, Ron D. Laboratory of Molecular Endocrinology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA. CHOP (GADD153) is a mammalian nuclear protein that dimerizes with members of the C/EBP family of transcriptional factors. Absent under normal conditions, CHOP is induced by the stress encountered during nutrient deprivation, the acute-phase response, and treatment of cells with certain toxins. The basic region of CHOP deviates considerably in sequence from that of other C/EBP proteins, and CHOP-C/EBP heterodimers are incapable of binding to a common class of C/EBP sites. With respect to such sites, CHOP serves as an inhibitor of the activity of C/EBP proteins. However, recent studies indicate that certain functions of CHOP, such as the induction of growth arrest by overexpression of the wild-type protein and oncogenic transformation by the TLS-CHOP fusion protein, require an intact basic region, suggesting that DNA binding by CHOP may be implicated in these activities. In this study an in vitro PCR-based selection assay was used to identify sequences bound by CHOP-C/EBP dimers. These sequences were found to contain a unique core element PuPuPuTGCAAT(A/C)CCC. Competition in DNA-binding assays, DNase 1 footprint analysis, and methylation interference demonstrate that the binding is sequence specific. Deletions in the basic region of CHOP lead to a loss of DNA binding, suggesting that CHOP participates in this process. Stress induction in NIH 3T3 cells leads to the appearance of CHOP-containing DNA-binding activity. CHOP is found to contain a transcriptional activation domain which is inducible by cellular stress, lending further support to the notion that the protein can function as a positively acting transcription factor. We conclude that CHOP may serve a dual role both as an inhibitor of the ability of C/EBP proteins to activate some target genes and as a direct activator of others. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8657121 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8674817 # MOTIF: [JP] MA0074.1 RXRA::VDR ---------------------------------------------- Mol Cell Endocrinol. 1995 Aug 30;113(1):89-98. Identification of DNA sequences that bind retinoid X receptor-1,25(OH)2D3-receptor heterodimers with high affinity. Colnot S, Lambert M, Blin C, Thomasset M, Perret C. INSERM U120, Hopital Robert Debre, Paris, France. Vitamin D3 receptors (VDR) bind as heterodimers with retinoid X receptors (RXR) to vitamin D response elements (VDRE) and transactivate gene expression in a 1,25(OH)2D3-dependent manner. These elements are tandem direct repeats (DRs) of the hexamer RGGTCA separated by three nucleotides (DR3). We determined whether this DR3 was the optimal and/or only recognition sequence, by PCR-mediated binding site selection with reticulocyte lysate-expressed hVDR and mRXRalpha, and a pool of random sequences. We derived a consensus binding site for RXR-VDR heterodimers, RGGTCANN RRGTTCAB, and analyzed 10 of the 45 sequences slected by EMSA, methylation interference and transfection experiments: all the sequences were specific and acted as positive VDREs; the underlined purine of the spacer interacted with the heterodimer; the mutation of the third T in the second motif to a G did not influence VDRE activity. Thus, the selectivity of vitamin D pathway involving heterodimerization rather than VDR-homodimerization is not due to internal sequence variations. Except for mouse osteopontin VDRE, the natural VDREs would be efficient, only when helped by adjacent sequences and/or transactivators other than VDR and RXR. PMID: 8674817 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16704374 # MOTIF: [JP] MA0136.1 ELF5 ---------------------------------------------- Biochem J. 2006 Sep 15;398(3):497-507. Determination of the consensus DNA-binding sequence and a transcriptional activation domain for ESE-2. Choi YS, Sinha S. Department of Biochemistry, State University of New York at Buffalo, Buffalo, NY 14214, USA. The ESE (epithelium-specific Ets) subfamily of Ets transcription factors plays an important role in regulating gene expression in a variety of epithelial cell types. Although ESE proteins have been shown to bind to regulatory elements of some epithelial genes, the optimal DNA-binding sequence has not been experimentally ascertained for any member of the ESE subfamily of transcription factors. This has made the identification and validation of their targets difficult. We are studying ESE-2 (Elf5), which is highly expressed in epithelial cells of many tissues including skin keratinocytes. Here, we identify the preferred DNA-binding site of ESE-2 by performing CASTing (cyclic amplification and selection of targets) experiments. Our analysis shows that the optimal ESE-2 consensus motif consists of a GGA core and an AT-rich 5'- and 3'-flanking sequences. Mutational and competition experiments demonstrate that the flanking sequences that confer high DNA-binding affinity for ESE-2 show considerable differences from the known consensus DNA-binding sites of other Ets proteins, thus reinforcing the idea that the flanking sequences may impart recognition specificity for Ets proteins. In addition, we have identified a novel isoform of murine ESE-2, ESE-2L, that is generated by use of a hitherto unreported new exon and an alternate promoter. Interestingly, transient transfection assays with an optimal ESE-2 responsive reporter show that both ESE-2 and ESE-2L are weak transactivators. However, similar studies utilizing GAL4 chimaeras of ESE-2 demonstrate that while the DNA-binding ETS (E twenty-six) domain functions as a repressor, the PNT (pointed domain) of ESE-2 can act as a potent transcriptional activation domain. This novel transactivating property of PNT is also shared by ESE-3, another ESE family member. Identification of the ESE-2 consensus site and characterization of the transcriptional activation properties of ESE-2 shed new light on its potential as a regulator of target genes. Publication Types: Research Support, N.I.H., Extramural PMID: 16704374 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16998473 # MOTIF: [JP] MA0264.1 ceh-22 ---------------------------------------------- Nat Biotechnol. 2006 Nov;24(11):1429-35. Epub 2006 Sep 24. Compact, universal DNA microarrays to comprehensively determine transcription-factor binding site specificities. Berger MF, Philippakis AA, Qureshi AM, He FS, Estep PW 3rd, Bulyk ML. Division of Genetics, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA. Transcription factors (TFs) interact with specific DNA regulatory sequences to control gene expression throughout myriad cellular processes. However, the DNA binding specificities of only a small fraction of TFs are sufficiently characterized to predict the sequences that they can and cannot bind. We present a maximally compact, synthetic DNA sequence design for protein binding microarray (PBM) experiments that represents all possible DNA sequence variants of a given length k (that is, all 'k-mers') on a single, universal microarray. We constructed such all k-mer microarrays covering all 10-base pair (bp) binding sites by converting high-density single-stranded oligonucleotide arrays to double-stranded (ds) DNA arrays. Using these microarrays we comprehensively determined the binding specificities over a full range of affinities for five TFs of different structural classes from yeast, worm, mouse and human. The unbiased coverage of all k-mers permits high-throughput interrogation of binding site preferences, including nucleotide interdependencies, at unprecedented resolution. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 16998473 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 11602361 # MOTIF: [JP] MA0135.1 Lhx3 ---------------------------------------------- Gene. 2001 Oct 17;277(1-2):239-50. Role of the LIM domains in DNA recognition by the Lhx3 neuroendocrine transcription factor. Bridwell JA, Price JR, Parker GE, McCutchan Schiller A, Sloop KW, Rhodes SJ. Department of Biology, Indiana University-Purdue University Indianapolis, 723 West Michigan Street, Indianapolis, IN 46202-5132, USA. LIM homeodomain transcription factors regulate many aspects of development in multicellular organisms. Such factors contain two LIM domains in their amino terminus and a DNA-binding homeodomain. To better understand the mechanism of gene regulation by these proteins, we studied the role of the LIM domains in DNA interaction by Lhx3, a protein that is essential for pituitary development and motor neuron specification in mammals. By site selection, we demonstrate that Lhx3 binds at high affinity to an AT-rich consensus DNA sequence that is similar to sequences located within the promoters of some pituitary hormone genes. The LIM domains reduce the affinity of DNA binding by Lhx3, but do not affect the specificity. Lhx3 preferentially binds to the consensus site as a monomer with minor groove contacts. The Lhx3 binding consensus site confers Lhx3-dependent transcriptional activation to heterologous promoters. Further, DNA molecules containing the consensus Lhx3 binding site are bent to similar angles in complexes containing either wild type Lhx3 or Lhx3 lacking LIM domains. These data are consistent with Lhx3 having the properties of an architectural transcription factor. We also propose that there are distinct classes of LIM homeodomain transcription factors in which the LIM domains play different roles in modulating interactions with DNA sites in target genes. Publication Types: Comparative Study Research Support, U.S. Gov't, Non-P.H.S. PMID: 11602361 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9443972 # MOTIF: [JP] MA0039.1 Klf4 ---------------------------------------------- Nucleic Acids Res. 1998 Feb 1;26(3):796-802. Identification of the DNA sequence that interacts with the gut-enriched Kruppel-like factor. Shields JM, Yang VW. Department of Medicine and Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. The gut-enriched Kruppel-like factor (GKLF) is a recently identified eukaryotic transcription factor that contains three C2H2zinc fingers. The amino acid sequence of the zinc finger portion of GKLF is closely related to several Kruppel proteins, including the lung Kruppel-like factor (LKLF), the erythroid Kruppel-like factor (EKLF) and the basic transcription element binding protein 2 (BTEB2). The DNA sequence to which GKLF binds has not been definitively established. In the present study we determined the DNA binding sequence of GKLF using highly purified recombinant GKLF in a target detection assay of an oligonucleotide library consisting of random sequences. Upon repeated rounds of selection and subsequent characterization of the selected sequences by base-specific mutagenesis a DNA with the sequence 5'-G/AG/AGGC/TGC/T-3' was found to contain the minimal essential binding site for GKLF. This sequence is present in the promoters of two previously characterized genes: the CACCC element of the beta-globin gene, which interacts with EKLF, and the basic transcription element (BTE) of the CYP1A1 gene, which interacts with Sp1 and several Sp1-like transcription factors. Moreover, the selected GKLF binding sequence was capable of mediating transactivation of a linked reporter gene by GKLF in co-transfection experiments. Our results establish GKLF as a sequence-specific transcription factor likely involved in regulation of expression of endogenous genes. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 9443972 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9826749 # MOTIF: [JP] MA0082.1 squamosa ---------------------------------------------- Nucleic Acids Res. 1998 Dec 1;26(23):5277-87. DNA binding and dimerisation determinants of Antirrhinum majus MADS-box transcription factors. West AG, Causier BE, Davies B, Sharrocks AD. Department of Biochemistry and Genetics, The Medical School, University of Newcastle upon Tyne, UK. Members of the MADS-box family of transcription factors are found in eukaryotes ranging from yeast to humans. In plants, MADS-box proteins regulate several developmental processes including flower, fruit and root development. We have investigated the DNA-binding mechanisms used by four such proteins in Antirrhinum majus, SQUA, PLE, DEF and GLO. SQUA differs from the characterised mammalian and yeast MADS-box proteins as it can efficiently bind two different classes of DNA-binding site. SQUA induces bending of these binding sites and the sequence of the site plays a role in determining the magnitude of these bends. Similarly, PLE and DEF/GLO induce DNA bending although the direction of the resulting bends differ. Finally, we demonstrate that the MADS-box and I-domains are sufficient for homodimer formation by SQUA. However, the K-box in SQUA can also act as an oligomerisation motif and in the full-length protein, the K-box plays a different role in mediating dimerisation in the context of SQUA homodimers or heterodimers with PLE. Together these results contribute significantly to our understanding of the function of SQUA and other plant MADS-box proteins at the molecular level. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 9826749 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10074718 # MOTIF: [JP] MA0020.1 Dof2 # MOTIF: [JP] MA0021.1 Dof3 # MOTIF: [JP] MA0053.1 MNB1A # MOTIF: [JP] MA0064.1 PBF ---------------------------------------------- Plant J. 1999 Jan;17(2):209-14. Diversity and similarity among recognition sequences of Dof transcription factors. Yanagisawa S, Schmidt RJ. Department of Life Sciences (Chemistry), Graduate School of Arts and Sciences, University of Tokyo, Japan. csyanag@komaba.ecc.u-tokyo.ac.jp Dof proteins are a family of transcription factors that share a unique DNA-binding domain. Dof proteins were found recently in association with diverse promoters of plant-specific genes, suggesting various roles of Dof proteins in plants. Through binding site selection experiments using randomly synthesized DNA, the recognition sequences of four maize Dof proteins were systematically analyzed. All selected oligonucleotides contained an AAAG sequence, suggesting that this sequence is the recognition core of Dof proteins. In fact, a single mutation in this sequence abolished binding of all four Dof proteins. Furthermore, the preference of each Dof protein for the sequence flanking the core motif was also analyzed using oligonucleotides containing a fixed AAAG and random flanking sequences. Similar, as well as distinct, flanking sequences were observed among the optimal binding sites. Changes in the flanking sequences did affect DNA-binding of Dof proteins. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 10074718 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17558387 # MOTIF: [JP] MA0137.1 STAT1 # MOTIF: [JP] MA0137.2 STAT1 ---------------------------------------------- Nat Methods. 2007 Aug;4(8):651-7. Epub 2007 Jun 11. Comment in: Nat Methods. 2007 Aug;4(8):613-4. Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencing. Robertson G, Hirst M, Bainbridge M, Bilenky M, Zhao Y, Zeng T, Euskirchen G, Bernier B, Varhol R, Delaney A, Thiessen N, Griffith OL, He A, Marra M, Snyder M, Jones S. British Columbia Cancer Agency Genome Sciences Centre, 675 West 10th Avenue, Vancouver, British Columbia V5Z 4S6, Canada. We developed a method, ChIP-sequencing (ChIP-seq), combining chromatin immunoprecipitation (ChIP) and massively parallel sequencing to identify mammalian DNA sequences bound by transcription factors in vivo. We used ChIP-seq to map STAT1 targets in interferon-gamma (IFN-gamma)-stimulated and unstimulated human HeLa S3 cells, and compared the method's performance to ChIP-PCR and to ChIP-chip for four chromosomes. By ChIP-seq, using 15.1 and 12.9 million uniquely mapped sequence reads, and an estimated false discovery rate of less than 0.001, we identified 41,582 and 11,004 putative STAT1-binding regions in stimulated and unstimulated cells, respectively. Of the 34 loci known to contain STAT1 interferon-responsive binding sites, ChIP-seq found 24 (71%). ChIP-seq targets were enriched in sequences similar to known STAT1 binding motifs. Comparisons with two ChIP-PCR data sets suggested that ChIP-seq sensitivity was between 70% and 92% and specificity was at least 95%. Publication Types: Research Support, Non-U.S. Gov't PMID: 17558387 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18842628 # MOTIF: [JP] MA0269.1 AFT1 # MOTIF: [JP] MA0273.1 ARO80 # MOTIF: [JP] MA0278.1 BAS1 # MOTIF: [JP] MA0296.1 FKH1 # MOTIF: [JP] MA0299.1 GAL4 # MOTIF: [JP] MA0303.1 GCN4 # MOTIF: [JP] MA0308.1 GSM1 # MOTIF: [JP] MA0336.1 MGA1 # MOTIF: [JP] MA0343.1 NDT80 # MOTIF: [JP] MA0345.1 NHP6A # MOTIF: [JP] MA0346.1 NHP6B # MOTIF: [JP] MA0347.1 NRG1 # MOTIF: [JP] MA0350.1 TOD6 # MOTIF: [JP] MA0351.1 DOT6 # MOTIF: [JP] MA0357.1 PHO4 # MOTIF: [JP] MA0376.1 RTG3 # MOTIF: [JP] MA0377.1 SFL1 # MOTIF: [JP] MA0378.1 SFP1 # MOTIF: [JP] MA0383.1 SMP1 # MOTIF: [JP] MA0386.1 TBP # MOTIF: [JP] MA0390.1 STB3 # MOTIF: [JP] MA0395.1 STP2 # MOTIF: [JP] MA0400.1 SUT2 # MOTIF: [JP] MA0413.1 USV1 # MOTIF: [JP] MA0415.1 YAP1 # MOTIF: [JP] MA0418.1 YAP6 # MOTIF: [JP] MA0435.1 YPR015C ---------------------------------------------- Nucleic Acids Res. 2009 Jan;37(Database issue):D77-82. Epub 2008 Oct 8. UniPROBE: an online database of protein binding microarray data on protein-DNA interactions. Newburger DE, Bulyk ML. Division of Genetics, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School and Harvard-MIT Division of Health Sciences and Technology, Harvard Medical School, Boston, MA 02115, USA. The UniPROBE (Universal PBM Resource for Oligonucleotide Binding Evaluation) database hosts data generated by universal protein binding microarray (PBM) technology on the in vitro DNA-binding specificities of proteins. This initial release of the UniPROBE database provides a centralized resource for accessing comprehensive PBM data on the preferences of proteins for all possible sequence variants ('words') of length k ('k-mers'), as well as position weight matrix (PWM) and graphical sequence logo representations of the k-mer data. In total, the database hosts DNA-binding data for over 175 nonredundant proteins from a diverse collection of organisms, including the prokaryote Vibrio harveyi, the eukaryotic malarial parasite Plasmodium falciparum, the parasitic Apicomplexan Cryptosporidium parvum, the yeast Saccharomyces cerevisiae, the worm Caenorhabditis elegans, mouse and human. Current web tools include a text-based search, a function for assessing motif similarity between user-entered data and database PWMs, and a function for locating putative binding sites along user-entered nucleotide sequences. The UniPROBE database is available at http://thebrain.bwh.harvard.edu/uniprobe/. Publication Types: Research Support, N.I.H., Extramural PMID: 18842628 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17210644 # MOTIF: [JP] POL011.1 XCPE1 ---------------------------------------------- Mol Cell Biol. 2007 Mar;27(5):1844-58. Epub 2007 Jan 8. The new core promoter element XCPE1 (X Core Promoter Element 1) directs activator-, mediator-, and TATA-binding protein-dependent but TFIID-independent RNA polymerase II transcription from TATA-less promoters. Tokusumi Y, Ma Y, Song X, Jacobson RH, Takada S. Department of Biochemistry and Molecular Biology, The University of Texas M. D. Anderson Cancer Center, Unit 1000, 1515 Holcombe Blvd., Houston, TX 77030, USA. The core promoter is a critical DNA element required for accurate transcription and regulation of transcription. Several core promoter elements have been previously identified in eukaryotes, but those cannot account for transcription from most RNA polymerase II-transcribed genes. Additional, as-yet-unidentified core promoter elements must be present in eukaryotic genomes. From extensive analyses of the hepatitis B virus X gene promoter, here we identify a new core promoter element, XCPE1 (the X gene core promoter element 1), that drives RNA polymerase II transcription. XCPE1 is located between nucleotides -8 and +2 relative to the transcriptional start site (+1) and has a consensus sequence of G/A/T-G/C-G-T/C-G-G-G/A-A-G/C(+1)-A/C. XCPE1 shows fairly weak transcriptional activity alone but exerts significant, specific promoter activity when accompanied by activator-binding sites. XCPE1 is also found in the core promoter regions of about 1% of human genes, particularly in poorly characterized TATA-less genes. Our in vitro transcription studies suggest that the XCPE1-driven transcription can be highly active in the absence of TFIID because it can utilize either free TBP or the complete TFIID complex. Our findings suggest the possibility of the existence of a TAF1 (TFIID)-independent transcriptional initiation mechanism that may be used by a category of TATA-less promoters in higher eukaryotes. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 17210644 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10561063 # MOTIF: [JP] MA0128.1 EmBP-1 # MOTIF: [JP] MA0129.1 TGA1A ---------------------------------------------- Plant Mol Biol. 1999 Sep;41(1):1-13. Bipartite determinants of DNA-binding specificity of plant basic leucine zipper proteins. Niu X, Renshaw-Gegg L, Miller L, Guiltinan MJ. Pioneer Hi-Bred International, Inc., Johnston, IA 50131, USA. The basic leucine zipper (bZIP) proteins are one of the largest and most conserved groups of eukaryotic transcription factors/repressors. Two major subgroups among the plant bZIP proteins have been identified as G-box (CCACGTGG) or C-box (TGACGTCA) binding proteins based on their DNA binding specificity and the amino acid sequences of their basic regions. We have investigated how plant bZIP proteins determine their DNA binding specificity by mutation of the basic domain of the G-box-binding protein EmBP-1. Four subregions of the EmBP-1 basic domain that differ from the C-box-binding protein TGA1a were substituted singly or in combination with the corresponding regions of TGA1a. DNA binding experiments with the mutant proteins demonstrated that binding specificity of plant bZIP proteins is determined independently by two regions, the core basic region and the hinge region. These two regions have an additive effect on DNA binding specificity. PCR-assisted binding-site selections using key mutants demonstrated that only G-box and C-box binding specificity can be generated by combinations of amino acids in the basic domains of EmBP-1 and TGA1a. These results suggest that factorial contributions of the amino acid residues in the basic domain combine to determine DNA-binding specificity of bZIP proteins. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 10561063 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10187832 # MOTIF: [JP] MA0115.1 NR1H2::RXRA ---------------------------------------------- J Biol Chem. 1999 Apr 9;274(15):10421-9. Identification of a novel DNA binding site for nuclear orphan receptor OR1. Feltkamp D, Wiebel FF, Alberti S, Gustafsson JA. Department of Biosciences at Novum, Karolinska Institute, Halsovagen 7, S-14157 Huddinge, Sweden. dorothee.feltkamp@cbt.ki.se The nuclear orphan receptor OR1 has been shown to bind as a heterodimer with retinoid X receptor (RXR) to direct repeat 4 (DR4) response elements. It remained unclear, however, whether this represents the only or the optimal binding site for this receptor. Therefore, we performed a DNA binding site selection assay that allows the identification of novel DNA binding sites for OR1 in an unbiased manner. While OR1 alone was not able to select a specific sequence from the pool of oligonucleotides, the OR1/RXR heterodimer selected a highly conserved DR1 element, termed DR1s, with two AGGTCA motifs spaced by one adenosine. The functional activity of the consensus binding site was verified in transient transfection assays and corroborated by in vitro studies. Based on the sequence of the consensus DR1s, we located putative natural binding sites in the 5'-promoter flanking regions of the rat S14 gene and the rat cholecystokinin type A receptor gene. Furthermore, we could show that although the OR1/RXR heterodimer has a distinct binding orientation on a DR4 element, it is able to bind in both orientations to the DR1s element. The OR1 paralog LXRalpha does not bind as a heterodimer with RXR to the DR1s element, indicating that these receptors, despite their homology, are involved in the regulation of different sets of genes. Publication Types: Research Support, Non-U.S. Gov't PMID: 10187832 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8496174 # MOTIF: [JP] MA0017.1 NR2F1 ---------------------------------------------- J Biol Chem. 1993 May 25;268(15):11125-33. Chicken ovalbumin upstream promoter-transcription factor (COUP-TF) represses transcription from the promoter of the gene for ornithine transcarbamylase in a manner antagonistic to hepatocyte nuclear factor-4 (HNF-4). Kimura A, Nishiyori A, Murakami T, Tsukamoto T, Hata S, Osumi T, Okamura R, Mori M, Takiguchi M. Department of Molecular Genetics, Kumamoto University School of Medicine, Japan. Chicken ovalbumin upstream promoter-transcription factor (COUP-TF) and hepatocyte nuclear factor-4 (HNF-4) are orphan members of the steroid/thyroid receptor superfamily and exhibit ubiquitous and liver-enriched tissue distribution, respectively. The gene for rat ornithine transcarbamylase (OTC), an ornithine cycle enzyme, is mainly expressed in the liver and is under the control of the promoter and the 11-kilobase upstream enhancer, both of which are liver-selective. Two sites of the promoter region and two sites of the enhancer region of the OTC gene, as well as the ovalbumin promoter site, were recognized by both HNF-4 and COUP-TF, showing that these two factors have closely related binding specificities. Since HNF-4 activated expression from the OTC promoter in cotransfection analysis, this factor appears to participate in liver-selective activation of the OTC gene. On the other hand, COUP-TF repressed the expression from the OTC promoter, whereas it activated expression from several other promoters. Therefore, COUP-TF plays a dual regulatory role depending on the promoter context. Repression of a tissue-specific promoter by a ubiquitous transactivator and derepression by a related tissue-enriched transactivator is potentially an important mechanism for tissue-specific activation of a gene. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8496174 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8052536 # MOTIF: [JP] MA0093.1 USF1 ---------------------------------------------- Nucleic Acids Res. 1994 Jul 25;22(14):2801-10. Base preferences for DNA binding by the bHLH-Zip protein USF: effects of MgCl2 on specificity and comparison with binding of Myc family members. Bendall AJ, Molloy PL. CSIRO Division of Biomolecular Engineering, North Ryde, NSW, Australia. Studies of the DNA binding specificity of transcription factors belonging to the basic helix-loop-helix (bHLH) family have identified the so-called E-box, CACGTG, as being a high affinity specific binding sequence for this class of DNA binding proteins. Binding sequences for HeLa USF were selected from an initially random population of 20 bp sequences, defining the optimum USF binding sequence as R-5Y-4C-3A-2C-1G+1T+2G+3R+4Y+5. The significance of the flanking bases was further demonstrated by showing that USF and the related proteins c-Myc and Max discriminate between CACGTG-type E-boxes and that the primary means of discrimination appears to be the identity of the nucleotide at +/- 4, the presence of a T at -4 being inhibitory to binding by Myc but not by USF or Max. This suggests one mechanism by which bHLH factors are partitioned between multiple potential binding sequences in the promoters and enhancers of viral and cellular genes. It was also demonstrated that MgCl2 has a significant influence on USF DNA binding specificity. A broader range of USF binding sites was selected in the absence of MgCl2, conforming to the altered half-site consensus gTGaY. Binding studies with specific oligonucleotides demonstrated significantly improved tolerance to sequence variation at positions 1, 4, and to a lesser extent 5, of the GTGRY consensus in the absence of MgCl2. The results indicate that Mg2+ ions have an integral role in the formation of the USF-DNA complex. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8052536 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10567552 # MOTIF: [JP] MA0068.1 Pax4 ---------------------------------------------- Mol Cell Biol. 1999 Dec;19(12):8272-80. Paired-homeodomain transcription factor PAX4 acts as a transcriptional repressor in early pancreatic development. Smith SB, Ee HC, Conners JR, German MS. Hormone Research Institute, University of California, San Francisco, San Francisco, California 94143-0534, USA. The paired-homeodomain transcription factor PAX4 is expressed in the developing pancreas and along with PAX6 is required for normal development of the endocrine cells. In the absence of PAX4, the numbers of insulin-producing beta cells and somatostatin-producing delta cells are drastically reduced, while the numbers of glucagon-producing alpha cells are increased. To gain insight into PAX4 function, we cloned a full-length Pax4 cDNA from a beta-cell cDNA library and identified a bipartite consensus DNA binding sequence consisting of a homeodomain binding site separated from a paired domain binding site by 15 nucleotides. The paired half of this consensus sequence has similarities to the PAX6 paired domain consensus binding site, and the two proteins bind to common sequences in several islet genes, although with different relative affinities. When expressed in an alpha-cell line, PAX4 represses transcription through the glucagon or insulin promoters or through an isolated PAX4 binding site. This repression is not simply due to competition with the PAX6 transcriptional activator for the same binding site, since PAX4 fused to the unrelated yeast GAL4 DNA binding domain also represses transcription through the GAL4 binding site in the alpha-cell line and to a lesser degree in beta-cell lines and NIH 3T3 cells. Repressor activity maps to more than one domain within the molecule, although the homeodomain and carboxyl terminus give the strongest repression. PAX4 transcriptional regulation apparently plays a role only early in islet development, since Pax4 mRNA as determined by reverse transcriptase PCR peaks at embryonic day 13.5 in the fetal mouse pancreas and is undetectable in adult islets. In summary, PAX4 can function as a transcriptional repressor and is expressed early in pancreatic development, which may allow it to suppress alpha-cell differentiation and permit beta-cell differentiation. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 10567552 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 17916232 # MOTIF: [JP] MA0018.2 CREB1 # MOTIF: [JP] MA0079.2 SP1 # MOTIF: [JP] MA0080.2 SPI1 # MOTIF: [JP] MA0099.2 AP1 # MOTIF: [JP] MA0150.1 NFE2L2 # MOTIF: [JP] MA0152.1 NFATC2 # MOTIF: [JP] MA0153.1 HNF1B # MOTIF: [JP] MA0154.1 EBF1 # MOTIF: [JP] MA0155.1 INSM1 # MOTIF: [JP] MA0156.1 FEV # MOTIF: [JP] MA0157.1 FOXO3 # MOTIF: [JP] MA0158.1 HOXA5 # MOTIF: [JP] MA0159.1 RXR::RAR_DR5 # MOTIF: [JP] MA0160.1 NR4A2 # MOTIF: [JP] MA0442.1 SOX10 ---------------------------------------------- Genome Biol. 2007;8(10):R207. PAZAR: a framework for collection and dissemination of cis-regulatory sequence annotation. Portales-Casamar E, Kirov S, Lim J, Lithwick S, Swanson MI, Ticoll A, Snoddy J, Wasserman WW. Centre for Molecular Medicine and Therapeutics, CFRI, University of British Columbia, Vancouver, BC, V5Z 4H4, Canada. elodie@cmmt.ubc.ca PAZAR is an open-access and open-source database of transcription factor and regulatory sequence annotation with associated web interface and programming tools for data submission and extraction. Curated boutique data collections can be maintained and disseminated through the unified schema of the mall-like PAZAR repository. The Pleiades Promoter Project collection of brain-linked regulatory sequences is introduced to demonstrate the depth of annotation possible within PAZAR. PAZAR, located at http://www.pazar.info, is open for business. Publication Types: Research Support, Non-U.S. Gov't PMID: 17916232 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7590239 # MOTIF: [JP] MA0085.1 Su(H) ---------------------------------------------- Genes Dev. 1995 Nov 1;9(21):2609-22. Suppressor of hairless directly activates transcription of enhancer of split complex genes in response to Notch receptor activity. Bailey AM, Posakony JW. Department of Biology, University of California San Diego, La Jolla 92093-0366, USA. We have investigated the functional relationships among three loci that are required for multiple alternative cell fate decisions during adult peripheral neurogenesis in Drosophila: Notch (N), which encodes a transmembrane receptor protein, Suppressor of Hairless [Su(H)], which encodes a DNA-binding transcription factor, and the Enhancer of split gene complex [E(spl)-C], which includes seven transcription units that encode basic helix-loop-helix (bHLH) repressor proteins. We describe several lines of evidence establishing that Su(H) directly activates transcription of E(spl)-C genes in response to N receptor activity. Expression of an activated form of the N receptor leads to elevated and ectopic E(spl)-C transcript accumulation and promoter activity in imaginal discs. We show that the proximal upstream regions of three E(spl)-C genes contain multiple specific binding sites for Su(H). The integrity of these sites, as well as Su(H) gene activity, are required not only for normal levels of expression of E(spl)-C genes in imaginal disc proneural clusters, but also for their transcriptional response to hyperactivity of the N receptor. Our results establish Su(H) as a direct regulatory link between N receptor activity and the expression of E(spl)-C genes, extending the known linear structure of the N cell-cell signaling pathway. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 7590239 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9774661 # MOTIF: [JP] MA0116.1 Zfp423 ---------------------------------------------- Mol Cell Biol. 1998 Nov;18(11):6447-56. Identification of DNA recognition sequences and protein interaction domains of the multiple-Zn-finger protein Roaz. Tsai RY, Reed RR. Howard Hughes Medical Institute, Department of Molecular Biology and Genetics and Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. Roaz, a rat C2H2 zinc finger protein, plays a role in the regulation of olfactory neuronal differentiation through its interaction with the Olf-1/EBF transcription factor family. An additional role for the Roaz/Olf-1/EBF heterodimeric protein is suggested by its ability to regulate gene activation at a distinct promoter lacking Olf-1/EBF-binding sites. Using an in vitro binding-site selection assay (Selex), we demonstrate that Roaz protein binds to novel inverted perfect or imperfect repeats of GCACCC separated by 2 bp. We show that Roaz is capable of binding to a canonical consensus recognition sequence with high affinity (Kd = 3 nM). Analysis of the structural requirement for protein dimerization and DNA binding by Roaz reveals the role of specific zinc finger motifs in the Roaz protein for homodimerization and heterodimerization with the Olf-1/EBF transcription factor. The DNA-binding domain of Roaz is mapped to the N-terminal 277 amino acids, containing the first seven zinc finger motifs, which confers weak monomeric binding to a single half site and a stronger dimeric binding to the inverted repeat in a binding-site-dependent manner. Full-length protein can form dimers on both the inverted repeat and direct repeat but not on a single half site. These findings support the role of the TFIIIA-type Zn fingers in both protein-protein interaction and protein-DNA interaction and suggest distinct functions for specific motifs in proteins with a large number of zinc finger structures. PMID: 9774661 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19632181 # MOTIF: [JP] PL0001.1 hlh-11 # MOTIF: [JP] PL0002.1 hlh-2::hlh-3 # MOTIF: [JP] PL0003.1 hlh-2::cnd-1 # MOTIF: [JP] PL0004.1 hlh-27 # MOTIF: [JP] PL0005.1 hlh-30 # MOTIF: [JP] PL0006.1 hlh-2::lin-32 # MOTIF: [JP] PL0007.1 mxl-3 # MOTIF: [JP] PL0008.1 hlh-29 # MOTIF: [JP] PL0009.1 hlh-26 # MOTIF: [JP] PL0010.1 hlh-2::hlh-19 # MOTIF: [JP] PL0011.1 hlh-2::hlh-4 # MOTIF: [JP] PL0012.1 hlh-2::hlh-8 # MOTIF: [JP] PL0013.1 hlh-2::hlh-15 # MOTIF: [JP] PL0014.1 mxl-1::mdl-1 # MOTIF: [JP] PL0015.1 hlh-2::hlh-14 # MOTIF: [JP] PL0016.1 ref-1 # MOTIF: [JP] PL0017.1 hlh-2::hlh-10 # MOTIF: [JP] PL0018.1 hlh-25 # MOTIF: [JP] PL0019.1 hlh-1 ---------------------------------------------- Cell. 2009 Jul 23;138(2):314-27. A multiparameter network reveals extensive divergence between C. elegans bHLH transcription factors. Grove CA, De Masi F, Barrasa MI, Newburger DE, Alkema MJ, Bulyk ML, Walhout AJ. Program in Gene Function and Expression and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA. Differences in expression, protein interactions, and DNA binding of paralogous transcription factors ("TF parameters") are thought to be important determinants of regulatory and biological specificity. However, both the extent of TF divergence and the relative contribution of individual TF parameters remain undetermined. We comprehensively identify dimerization partners, spatiotemporal expression patterns, and DNA-binding specificities for the C. elegans bHLH family of TFs, and model these data into an integrated network. This network displays both specificity and promiscuity, as some bHLH proteins, DNA sequences, and tissues are highly connected, whereas others are not. By comparing all bHLH TFs, we find extensive divergence and that all three parameters contribute equally to bHLH divergence. Our approach provides a framework for examining divergence for other protein families in C. elegans and in other complex multicellular organisms, including humans. Cross-species comparisons of integrated networks may provide further insights into molecular features underlying protein family evolution. For a video summary of this article, see the PaperFlick file available with the online Supplemental Data. PMID: 19632181 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1673656 # MOTIF: [JP] MA0094.1 Ubx ---------------------------------------------- EMBO J. 1991 May;10(5):1179-86. Optimal DNA sequence recognition by the Ultrabithorax homeodomain of Drosophila. Ekker SC, Young KE, von Kessler DP, Beachy PA. Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205. The 61 amino acid homeodomain is conserved among members of a family of eukaryotic DNA-binding proteins that play regulatory roles in transcription and in development. We have refined a rapid method for determining optimal DNA binding sites and have applied it to a 72 amino acid peptide containing the homeodomain of the Ultrabithorax (Ubx) homeotic gene of Drosophila. The site (5'-TTAATGG-3') is tightly bound (KD approximately 7 x 10(-11) M) by the Ubx homeodomain peptide; the four central TAAT bases of this sequence play a primary role in determining the affinity of binding, with significant secondary contributions deriving from the flanking bases. Although previously defined genomic sites contain multiple TAAT sequences with flanking bases distinct from those in the optimal binding site, we have found a new binding site with seven near-perfect repeats of the optimal sequence; this site is located in the promoter region of decapentaplegic, a probable Ubx regulatory target. The presence of a TAAT motif in the binding sites for most other homeodomain proteins suggests the existence of a conserved mechanism for recognition of this core sequence, with further specificity conferred by interactions with bases flanking this core. Publication Types: Research Support, Non-U.S. Gov't PMID: 1673656 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15177025 # MOTIF: [JP] MA0263.1 ttx-3::ceh-10 ---------------------------------------------- Dev Cell. 2004 Jun;6(6):757-70. Genomic cis-regulatory architecture and trans-acting regulators of a single interneuron-specific gene battery in C. elegans. Wenick AS, Hobert O. Department of Biochemistry and Molecular Biophysics, Center for Neurobiology and Behavior, Columbia University Medical Center, 701 West 168th Street, New York, NY 10032, USA. Gene batteries are sets of coregulated genes with common cis-regulatory elements that define the differentiated state of a cell. The nature of gene batteries for individual neuronal cellular subtypes and their linked cis-regulatory elements is poorly defined. Through molecular dissection of the highly modular cis-regulatory architecture of individual neuronally expressed genes, we have defined a conserved 16 bp cis-regulatory motif that drives gene expression in a single interneuron subtype, termed AIY, in the nematode Caenorhabditis elegans. This motif is bound and activated by the Paired- and LIM-type homeodomain proteins CEH-10 and TTX-3. Using genome-wide phylogenetic footprinting, we delineated the location, distribution, and evolution of AIY-specific cis-regulatory elements throughout the genome and thereby defined a large battery of AIY-expressed genes, all of which represent direct Paired/LIM homeodomain target genes. The identity of these homeodomain targets provides novel insights into the biology of the AIY interneuron. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 15177025 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1280827 # MOTIF: [JP] MA0016.1 usp ---------------------------------------------- Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11503-7. DNA binding and heteromerization of the Drosophila transcription factor chorion factor 1/ultraspiracle. Christianson AM, King DL, Hatzivassiliou E, Casas JE, Hallenbeck PL, Nikodem VM, Mitsialis SA, Kafatos FC. Department of Cellular and Developmental Biology, Harvard University, Cambridge, MA 02138. The Drosophila chorion factor 1/ultraspiracle (CF1/USP) transcription factor, a homologue of the retinoid X receptor, is a developmentally important member of the family of nuclear (steroid) hormone receptors. Using newly developed monoclonal antibodies and a full-length bacterially produced protein, we have studied in detail the in vitro DNA-binding properties of this factor and aspects of its distribution in vivo. During oogenesis, CF1/USP is present both in germline cells and in the somatic follicular epithelium. We have determined the optimal binding site of partially purified bacterially produced CF1/USP by an in vitro selection procedure and also have characterized its binding to the follicular-specific chorion s15 promoter. In vitro this bacterially produced factor is unusual in binding to a single element ("half-site"); simultaneous but noncoordinate binding to a second half-site is possible if these repeated elements are organized in direct orientation and spaced adequately. However, the factor interacts synergistically with several other nuclear hormone receptors: notably, it can form in vitro heteromers with mammalian thyroid and retinoic acid receptors, binding to two half-sites that are organized in either direct or inverted orientation. In vivo the factor most probably functions as a heterodimer, but its partner(s) remains to be determined. Publication Types: Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S. PMID: 1280827 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8816445 # MOTIF: [JP] MA0073.1 RREB1 ---------------------------------------------- Mol Cell Biol. 1996 Oct;16(10):5335-45. RREB-1, a novel zinc finger protein, is involved in the differentiation response to Ras in human medullary thyroid carcinomas. Thiagalingam A, De Bustros A, Borges M, Jasti R, Compton D, Diamond L, Mabry M, Ball DW, Baylin SB, Nelkin BD. Oncology Center, Johns Hopkins Medical Institutions, Baltimore, Maryland 21231, USA. An activated ras oncogene induces a program of differentiation in the human medullary thyroid cancer cell line TT. This differentiation process is accompanied by a marked increase in the transcription of the human calcitonin (CT) gene. We have localized a unique Ras-responsive transcriptional element (RRE) in the CT gene promoter. DNase I protection indicates two domains of protein-DNA interaction, and each domain separately can confer Ras-mediated transcriptional inducibility. This bipartite RRE was also found to be Raf responsive. By affinity screening, we have cloned a cDNA coding for a zinc finger transcription factor (RREB-1) that binds to the distal RRE. The consensus binding site for this factor is CCCCAAACCACCCC. RREB-1 is expressed ubiquitously in human tissues outside the adult brain. Overexpression of RREB-1 protein in TT cells confers the ability to mediate increased transactivation of the CT gene promoter-reporter construct during Ras- or Raf-induced differentiation. These data suggest that RREB-1 may play a role in Ras and Raf signal transduction in medullary thyroid cancer and other cells. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8816445 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12385991 # MOTIF: [JP] MA0114.1 HNF4A ---------------------------------------------- Bioinformatics. 2002;18 Suppl 2:S100-9. Identifying transcription factor binding sites through Markov chain optimization. Ellrott K, Yang C, Sladek FM, Jiang T. Department of Computer Science, University of California, Riverside, 92521, USA. Even though every cell in an organism contains the same genetic material, each cell does not express the same cohort of genes. Therefore, one of the major problems facing genomic research today is to determine not only which genes are differentially expressed and under what conditions, but also how the expression of those genes is regulated. The first step in determining differential gene expression is the binding of sequence-specific DNA binding proteins (i.e. transcription factors) to regulatory regions of the genes (i.e. promoters and enhancers). An important aspect to understanding how a given transcription factor functions is to know the entire gamut of binding sites and subsequently potential target genes that the factor may bind/regulate. In this study, we have developed a computer algorithm to scan genomic databases for transcription factor binding sites, based on a novel Markov chain optimization method, and used it to scan the human genome for sites that bind to hepatocyte nuclear factor 4 alpha (HNF4alpha). A list of 71 known HNF4alpha binding sites from the literature were used to train our Markov chain model. By looking at the window of 600 nucleotides around the transcription start site of each confirmed gene on the human genome, we identified 849 sites with varying binding potential and experimentally tested 109 of those sites for binding to HNF4alpha. Our results show that the program was very successful in identifying 77 new HNF4alpha binding sites with varying binding affinities (i.e. a 71% success rate). Therefore, this computational method for searching genomic databases for potential transcription factor binding sites is a powerful tool for investigating mechanisms of differential gene regulation. Publication Types: Comparative Study Evaluation Studies Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S. PMID: 12385991 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9047360 # MOTIF: [JP] MA0046.1 HNF1A ---------------------------------------------- J Mol Biol. 1997 Feb 21;266(2):231-45. Analysis of the distribution of binding sites for a tissue-specific transcription factor in the vertebrate genome. Tronche F, Ringeisen F, Blumenfeld M, Yaniv M, Pontoglio M. Unite des virus Oncogenes, URA 1644 du CNRS, Departement des Biotechnologies, Institut Pasteur, Paris, France. Hepatocyte nuclear factor 1 (HNF1) is a dimeric homeoprotein expressed in hepatocytes and in a few other epithelial cells where it helps regulate the expression of a specific subset of genes. In an attempt to identify novel target genes for HNF1 and to assess the distribution of its target sites within the vertebrate genome, we performed a computer-assisted search within the available databases using a weighted matrix. Several hundred potential target sequences were identified within the GenBank and EMBL data banks. DNA binding assays demonstrated that more than 95%, of the new sites tested (52 sites among 54) bound HNF1. Surprisingly many HNF1 target sites were found in genes that are transcribed in cell types that do not contain the protein. On the other hand these sites are 2.5 to five times more frequent in hepatic genes than expected. It seems that the presence of HNF1 sites in liver-specific genes was favoured, but that no counter-selection occurred within the rest of the genome. HNF1 binding sites in liver genes are more often associated in clusters with sites for other transcription factors and the enrichment is more pronounced in promoter regions. We identified more than 100 liver specific genes that are potentially regulated by HNF1. Publication Types: Research Support, Non-U.S. Gov't PMID: 9047360 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8264613 # MOTIF: [JP] MA0018.1 CREB1 ---------------------------------------------- Mol Cell Biol. 1994 Jan;14(1):456-62. In vitro selection of DNA elements highly responsive to the human T-cell lymphotropic virus type I transcriptional activator, Tax. Paca-Uccaralertkun S, Zhao LJ, Adya N, Cross JV, Cullen BR, Boros IM, Giam CZ. Department of Medicine, Case Western Reserve University, Cleveland, Ohio 44106. The human T-cell lymphotropic virus type I (HTLV-I) transactivator, Tax, the ubiquitous transcriptional factor cyclic AMP (cAMP) response element-binding protein (CREB protein), and the 21-bp repeats in the HTLV-I transcriptional enhancer form a ternary nucleoprotein complex (L. J. Zhao and C. Z. Giam, Proc. Natl. Acad. Sci. USA 89:7070-7074, 1992). Using an antibody directed against the COOH-terminal region of Tax along with purified Tax and CREB proteins, we selected DNA elements bound specifically by the Tax-CREB complex in vitro. Two distinct but related groups of sequences containing the cAMP response element (CRE) flanked by long runs of G and C residues in the 5' and 3' regions, respectively, were preferentially recognized by Tax-CREB. In contrast, CREB alone binds only to CRE motifs (GNTGACG[T/C]) without neighboring G- or C-rich sequences. The Tax-CREB-selected sequences bear a striking resemblance to the 5' or 3' two-thirds of the HTLV-I 21-bp repeats and are highly inducible by Tax. Gel electrophoretic mobility shift assays, DNA transfection, and DNase I footprinting analyses indicated that the G- and C-rich sequences flanking the CRE motif are crucial for Tax-CREB-DNA ternary complex assembly and Tax transactivation but are not in direct contact with the Tax-CREB complex. These data show that Tax recruits CREB to form a multiprotein complex that specifically recognizes the viral 21-bp repeats. The expanded DNA binding specificity of Tax-CREB and the obligatory role the ternary Tax-CREB-DNA complex plays in transactivation reveal a novel mechanism for regulating the transcriptional activity of leucine zipper proteins like CREB. Publication Types: In Vitro Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8264613 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18585359 # MOTIF: [JP] PH0001.1 Alx3 # MOTIF: [JP] PH0002.1 Alx4 # MOTIF: [JP] PH0003.1 Arx # MOTIF: [JP] PH0004.1 Nkx3-2 # MOTIF: [JP] PH0005.1 Barhl1 # MOTIF: [JP] PH0006.1 Barhl2 # MOTIF: [JP] PH0007.1 Barx1 # MOTIF: [JP] PH0008.1 Barx2 # MOTIF: [JP] PH0009.1 Bsx # MOTIF: [JP] PH0010.1 Alx1_1 # MOTIF: [JP] PH0011.1 Alx1_2 # MOTIF: [JP] PH0012.1 Cdx1 # MOTIF: [JP] PH0013.1 Cdx2 # MOTIF: [JP] PH0014.1 Cphx # MOTIF: [JP] PH0015.1 Crx # MOTIF: [JP] PH0016.1 Cux1_1 # MOTIF: [JP] PH0017.1 Cux1_2 # MOTIF: [JP] PH0018.1 Dbx1 # MOTIF: [JP] PH0019.1 Dbx2 # MOTIF: [JP] PH0020.1 Dlx1 # MOTIF: [JP] PH0021.1 Dlx2 # MOTIF: [JP] PH0022.1 Dlx3 # MOTIF: [JP] PH0023.1 Dlx4 # MOTIF: [JP] PH0024.1 Dlx5 # MOTIF: [JP] PH0025.1 Dmbx1 # MOTIF: [JP] PH0026.1 Duxbl # MOTIF: [JP] PH0027.1 Emx2 # MOTIF: [JP] PH0028.1 En1 # MOTIF: [JP] PH0029.1 En2 # MOTIF: [JP] PH0030.1 Esx1 # MOTIF: [JP] PH0031.1 Evx1 # MOTIF: [JP] PH0032.1 Evx2 # MOTIF: [JP] PH0033.1 Gbx1 # MOTIF: [JP] PH0034.1 Gbx2 # MOTIF: [JP] PH0035.1 Gsc # MOTIF: [JP] PH0036.1 Gsx2 # MOTIF: [JP] PH0037.1 Hdx # MOTIF: [JP] PH0038.1 Hlx # MOTIF: [JP] PH0039.1 Mnx1 # MOTIF: [JP] PH0040.1 Hmbox1 # MOTIF: [JP] PH0041.1 Hmx1 # MOTIF: [JP] PH0042.1 Hmx2 # MOTIF: [JP] PH0043.1 Hmx3 # MOTIF: [JP] PH0044.1 Homez # MOTIF: [JP] PH0045.1 Hoxa1 # MOTIF: [JP] PH0046.1 Hoxa10 # MOTIF: [JP] PH0047.1 Hoxa11 # MOTIF: [JP] PH0048.1 Hoxa13 # MOTIF: [JP] PH0049.1 Hoxa2 # MOTIF: [JP] PH0050.1 Hoxa3 # MOTIF: [JP] PH0051.1 Hoxa4 # MOTIF: [JP] PH0052.1 Hoxa5 # MOTIF: [JP] PH0053.1 Hoxa6 # MOTIF: [JP] PH0054.1 Hoxa7_1 # MOTIF: [JP] PH0055.1 Hoxa7_2 # MOTIF: [JP] PH0056.1 Hoxa9 # MOTIF: [JP] PH0057.1 Hoxb13 # MOTIF: [JP] PH0058.1 Hoxb3 # MOTIF: [JP] PH0059.1 Hoxb4 # MOTIF: [JP] PH0060.1 Hoxb5 # MOTIF: [JP] PH0061.1 Hoxb6 # MOTIF: [JP] PH0062.1 Hoxb7 # MOTIF: [JP] PH0063.1 Hoxb8 # MOTIF: [JP] PH0064.1 Hoxb9 # MOTIF: [JP] PH0065.1 Hoxc10 # MOTIF: [JP] PH0066.1 Hoxc11 # MOTIF: [JP] PH0067.1 Hoxc12 # MOTIF: [JP] PH0068.1 Hoxc13 # MOTIF: [JP] PH0069.1 Hoxc4 # MOTIF: [JP] PH0070.1 Hoxc5 # MOTIF: [JP] PH0071.1 Hoxc6 # MOTIF: [JP] PH0072.1 Hoxc8 # MOTIF: [JP] PH0073.1 Hoxc9 # MOTIF: [JP] PH0074.1 Hoxd1 # MOTIF: [JP] PH0075.1 Hoxd10 # MOTIF: [JP] PH0076.1 Hoxd11 # MOTIF: [JP] PH0077.1 Hoxd12 # MOTIF: [JP] PH0078.1 Hoxd13 # MOTIF: [JP] PH0079.1 Hoxd3 # MOTIF: [JP] PH0080.1 Hoxd8 # MOTIF: [JP] PH0081.1 Pdx1 # MOTIF: [JP] PH0082.1 Irx2 # MOTIF: [JP] PH0083.1 Irx3_1 # MOTIF: [JP] PH0084.1 Irx3_2 # MOTIF: [JP] PH0085.1 Irx4 # MOTIF: [JP] PH0086.1 Irx5 # MOTIF: [JP] PH0087.1 Irx6 # MOTIF: [JP] PH0088.1 Isl2 # MOTIF: [JP] PH0089.1 Isx # MOTIF: [JP] PH0090.1 Lbx2 # MOTIF: [JP] PH0091.1 Lhx1 # MOTIF: [JP] PH0092.1 Lhx2 # MOTIF: [JP] PH0093.1 Lhx3 # MOTIF: [JP] PH0094.1 Lhx4 # MOTIF: [JP] PH0095.1 Lhx5 # MOTIF: [JP] PH0096.1 Lhx6_1 # MOTIF: [JP] PH0097.1 Lhx6_2 # MOTIF: [JP] PH0098.1 Lhx8 # MOTIF: [JP] PH0099.1 Lhx9 # MOTIF: [JP] PH0100.1 Lmx1a # MOTIF: [JP] PH0101.1 Lmx1b # MOTIF: [JP] PH0102.1 Meis1 # MOTIF: [JP] PH0103.1 Meox1 # MOTIF: [JP] PH0104.1 Meis2 # MOTIF: [JP] PH0105.1 Meis3 # MOTIF: [JP] PH0106.1 Msx1 # MOTIF: [JP] PH0107.1 Msx2 # MOTIF: [JP] PH0108.1 Msx3 # MOTIF: [JP] PH0109.1 Nkx1-1 # MOTIF: [JP] PH0110.1 Nkx1-2 # MOTIF: [JP] PH0111.1 Nkx2-2 # MOTIF: [JP] PH0112.1 Nkx2-3 # MOTIF: [JP] PH0113.1 Nkx2-4 # MOTIF: [JP] PH0114.1 Nkx2-5 # MOTIF: [JP] PH0115.1 Nkx2-6 # MOTIF: [JP] PH0116.1 Nkx2-9 # MOTIF: [JP] PH0117.1 Nkx3-1 # MOTIF: [JP] PH0118.1 Nkx6-1_1 # MOTIF: [JP] PH0119.1 Nkx6-1_2 # MOTIF: [JP] PH0120.1 Nkx6-3 # MOTIF: [JP] PH0121.1 Obox1 # MOTIF: [JP] PH0122.1 Obox2 # MOTIF: [JP] PH0123.1 Obox3 # MOTIF: [JP] PH0124.1 Obox5_1 # MOTIF: [JP] PH0125.1 Obox5_2 # MOTIF: [JP] PH0126.1 Obox6 # MOTIF: [JP] PH0127.1 Nobox # MOTIF: [JP] PH0128.1 Otp # MOTIF: [JP] PH0129.1 Otx1 # MOTIF: [JP] PH0130.1 Otx2 # MOTIF: [JP] PH0131.1 Pax4 # MOTIF: [JP] PH0132.1 Pax6 # MOTIF: [JP] PH0133.1 Pax7 # MOTIF: [JP] PH0134.1 Pbx1 # MOTIF: [JP] PH0135.1 Phox2a # MOTIF: [JP] PH0136.1 Phox2b # MOTIF: [JP] PH0137.1 Pitx1 # MOTIF: [JP] PH0138.1 Pitx2 # MOTIF: [JP] PH0139.1 Pitx3 # MOTIF: [JP] PH0140.1 Pknox1 # MOTIF: [JP] PH0141.1 Pknox2 # MOTIF: [JP] PH0142.1 Pou1f1 # MOTIF: [JP] PH0143.1 Pou2f1 # MOTIF: [JP] PH0144.1 Pou2f2 # MOTIF: [JP] PH0145.1 Pou2f3 # MOTIF: [JP] PH0146.1 Pou3f1 # MOTIF: [JP] PH0147.1 Pou3f2 # MOTIF: [JP] PH0148.1 Pou3f3 # MOTIF: [JP] PH0149.1 Pou3f4 # MOTIF: [JP] PH0150.1 Pou4f3 # MOTIF: [JP] PH0151.1 Pou6f1_1 # MOTIF: [JP] PH0152.1 Pou6f1_2 # MOTIF: [JP] PH0153.1 Prop1 # MOTIF: [JP] PH0154.1 Prrx1 # MOTIF: [JP] PH0155.1 Prrx2 # MOTIF: [JP] PH0156.1 Rax # MOTIF: [JP] PH0157.1 Rhox11_1 # MOTIF: [JP] PH0158.1 Rhox11_2 # MOTIF: [JP] PH0159.1 Rhox6 # MOTIF: [JP] PH0160.1 Shox2 # MOTIF: [JP] PH0161.1 Six1 # MOTIF: [JP] PH0162.1 Six2 # MOTIF: [JP] PH0163.1 Six3 # MOTIF: [JP] PH0164.1 Six4 # MOTIF: [JP] PH0165.1 Six6_1 # MOTIF: [JP] PH0166.1 Six6_2 # MOTIF: [JP] PH0167.1 Tcf1 # MOTIF: [JP] PH0168.1 Hnf1b # MOTIF: [JP] PH0169.1 Tgif1 # MOTIF: [JP] PH0170.1 Tgif2 # MOTIF: [JP] PH0171.1 Nkx2-1 # MOTIF: [JP] PH0172.1 Tlx2 # MOTIF: [JP] PH0173.1 Uncx # MOTIF: [JP] PH0174.1 Vax1 # MOTIF: [JP] PH0175.1 Vax2 # MOTIF: [JP] PH0176.1 Vsx1 ---------------------------------------------- Cell. 2008 Jun 27;133(7):1266-76. Comment in: Cell. 2008 Jun 27;133(7):1133-5. Variation in homeodomain DNA binding revealed by high-resolution analysis of sequence preferences. Berger MF, Badis G, Gehrke AR, Talukder S, Philippakis AA, Pena-Castillo L, Alleyne TM, Mnaimneh S, Botvinnik OB, Chan ET, Khalid F, Zhang W, Newburger D, Jaeger SA, Morris QD, Bulyk ML, Hughes TR. Division of Genetics, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA. Most homeodomains are unique within a genome, yet many are highly conserved across vast evolutionary distances, implying strong selection on their precise DNA-binding specificities. We determined the binding preferences of the majority (168) of mouse homeodomains to all possible 8-base sequences, revealing rich and complex patterns of sequence specificity and showing that there are at least 65 distinct homeodomain DNA-binding activities. We developed a computational system that successfully predicts binding sites for homeodomain proteins as distant from mouse as Drosophila and C. elegans, and we infer full 8-mer binding profiles for the majority of known animal homeodomains. Our results provide an unprecedented level of resolution in the analysis of this simple domain structure and suggest that variation in sequence recognition may be a factor in its functional diversity and evolutionary success. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. PMID: 18585359 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8065331 # MOTIF: [JP] MA0043.1 HLF ---------------------------------------------- Mol Cell Biol. 1994 Sep;14(9):5986-96. DNA-binding and transcriptional regulatory properties of hepatic leukemia factor (HLF) and the t(17;19) acute lymphoblastic leukemia chimera E2A-HLF. Hunger SP, Brown R, Cleary ML. Department of Pathology, Stanford University School of Medicine, California 94305. The t(17;19) translocation in acute lymphoblastic leukemias results in creation of E2A-hepatic leukemia factor (HLF) chimeric proteins that contain the DNA-binding and protein dimerization domains of the basic leucine zipper (bZIP) protein HLF fused to a portion of E2A proteins with transcriptional activation properties. An in vitro binding site selection procedure was used to determine DNA sequences preferentially bound by wild-type HLF and chimeric E2A-HLF proteins isolated from various t(17;19)-bearing leukemias. All were found to selectively bind the consensus sequence 5'-GTTACGTAAT-3' with high affinity. Wild-type and chimeric HLF proteins also bound closely related sites identified previously for bZIP proteins of both the proline- and acidic amino acid-rich (PAR) and C/EBP subfamilies; however, E2A-HLF proteins were significantly less tolerant of certain deviations from the HLF consensus binding site. These differences were directly attributable to loss of an HLF ancillary DNA-binding domain in all E2A-HLF chimeras and were further exacerbated by a zipper mutation in one isolate. Both wild-type and chimeric HLF proteins displayed transcriptional activator properties in lymphoid and nonlymphoid cells on reporter genes containing HLF or C/EBP consensus binding sites. But on reporter genes with nonoptimal binding sites, their transcriptional properties diverged and E2A-HLF competitively inhibited activation by wild-type PAR proteins. These findings establish a spectrum of binding site-specific transcriptional properties for E2A-HLF which may preferentially activate expression of select subordinate genes as a homodimer and potentially antagonize expression of others through heteromeric interactions. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8065331 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 12368505 # MOTIF: [JP] MA0123.1 abi4 ---------------------------------------------- Plant Cell. 2002 Oct;14(10):2565-75. Maize ABI4 binds coupling element1 in abscisic acid and sugar response genes. Niu X, Helentjaris T, Bate NJ. Agronomic Traits, Trait and Technology Development, Pioneer Hi-Bred International, Johnston, IA 50131-0552, USA. Significant progress has been made in elucidating the mechanism of abscisic acid (ABA)-regulated gene expression, including the characterization of an ABA-responsive element (ABRE), which is regulated by basic domain/Leu zipper transcription factors. In addition to the ABRE, a coupling element (CE1) has been demonstrated to be involved in ABA-induced expression. However, a trans factor that interacts with CE1 has yet to be characterized. We report the isolation of a seed-specific maize ABI4 homolog and demonstrate, using a PCR-based in vitro selection procedure, that the maize ABI4 protein binds to the CE-1 like sequence CACCG. Using electrophoretic mobility shift assays, we demonstrate that recombinant ZmABI4 protein binds to the CE1 element in a number of ABA-related genes. ZmABI4 also binds to the promoter of the sugar-responsive ADH1 gene, demonstrating the ability of this protein to regulate both ABA- and sugar-regulated pathways. ZmABI4 complements Arabidopsis ABI4 function, because abi4 mutant plants transformed with the ZmABI4 gene have an ABA- and sugar-sensitive phenotype. Identification of the maize ABI4 ortholog and the demonstration of its binding to a known ABA response element provide a link between ABA-mediated kernel development and the regulation of ABA response genes. Publication Types: Comparative Study PMID: 12368505 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16230532 # MOTIF: [JP] POL007.1 BREd ---------------------------------------------- Genes Dev. 2005 Oct 15;19(20):2418-23. A core promoter element downstream of the TATA box that is recognized by TFIIB. Deng W, Roberts SG. Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, United Kingdom. We have defined a core promoter element downstream of the TATA box that is recognized by TFIIB. This involves a DNA-binding domain in TFIIB that is distinct from the helix-turn-helix motif (which recognizes an element upstream of the TATA box). The TFIIB recognition element we describe regulates transcription in a manner that is promoter context-dependent, particularly with respect to the TFIIB recognition element that is located upstream of the TATA box. Thus TFIIB can recognize two distinct sequence elements that flank the TATA box, employing independent DNA-binding motifs and cooperating in the regulation of transcription. Publication Types: Research Support, Non-U.S. Gov't PMID: 16230532 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8265351 # MOTIF: [JP] MA0058.1 MAX # MOTIF: [JP] MA0059.1 MYC::MAX ---------------------------------------------- Nucleic Acids Res. 1993 Nov 25;21(23):5372-6. Distinct DNA binding preferences for the c-Myc/Max and Max/Max dimers. Solomon DL, Amati B, Land H. Growth Control and Development Laboratory, Imperial Cancer Research Fund, London, UK. The transcription factor c-Myc and its dimerisation partner Max are members of the basic/helix-loop-helix/leucine-zipper (bHLH-Z) family and bind to the DNA core sequence CACGTG. Using a site-selection protocol, we determined the complete 12 base pair consensus binding sites of c-Myc/Max (RACCACGTGGTY) and Max/Max (RANCACGTGNTY) dimers. We find that the c-Myc/Max dimer fails to bind the core when it is flanked by a 5'T or a 3'A, while the Max/Max dimer readily binds such sequences. Furthermore we show that inappropriate flanking sequences preclude transactivation by c-Myc in vivo. In conclusion, Max/Max dimers are less discriminatory than c-Myc/Max and may regulate other genes in addition to c-Myc/Max targets. PMID: 8265351 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8344258 # MOTIF: [JP] MA0009.1 T ---------------------------------------------- EMBO J. 1993 Aug;12(8):3211-20. Erratum in: EMBO J. 1993 Dec;12(12):4898-9. The Brachyury gene encodes a novel DNA binding protein. Kispert A, Herrmann BG. Max-Planck-Institut fur Entwicklungsbiologie, Abt. Biochemie, Tubingen, Germany. Brachyury (T) mutant embryos are deficient in mesoderm formation and do not complete axial development. The notochord is most strongly affected. The T gene is expressed transiently in primitive streak-derived nascent and migrating mesoderm cells and continuously in the notochord. Ectopic expression of T protein in the animal cap of Xenopus embryos results in ectopic mesoderm formation. The T protein is located in the nucleus. These and other data suggested that the T gene might be involved in the control of transcriptional regulation. In an attempt to demonstrate specific DNA binding of the T protein we have identified a consensus sequence among DNA fragments selected from a mixture of random oligomers. Under our experimental conditions T protein binds as a monomer to DNA. This property resides in the N-terminal domain of 229 amino acid residues which is strongly conserved between the mouse protein, and its Xenopus and zebrafish homologues. The latter proteins also recognize the consensus DNA binding site. We suggest that the T protein is involved in the control of genes required for mesoderm formation, and for the differentiation and function of chorda mesoderm. Publication Types: Research Support, Non-U.S. Gov't PMID: 8344258 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1582412 # MOTIF: [JP] MA0022.1 dl_1 # MOTIF: [JP] MA0023.1 dl_2 ---------------------------------------------- EMBO J. 1992 May;11(5):1837-42. The same dorsal binding site mediates both activation and repression in a context-dependent manner. Pan D, Courey AJ. Department of Chemistry and Biochemistry, University of California, Los Angeles 90024-1569. Like many DNA binding transcription factors, the Drosophila morphogen encoded by dorsal can both stimulate and repress promoter activity. In particular, this factor activates twist and represses zerknullt on the ventral side of the early embryo. We find that when multiple copies of a dorsal binding site from the zerknullt ventral repressor element are fused to a heterologous basal promoter, the resulting construct is activated by dorsal to give a ventral specific expression pattern. Thus, the ability of a dorsal binding site to mediate repression rather than activation is not an intrinsic property of the site, but depends upon its context. We also show that a hybrid promoter containing both the zerknullt ventral repressor element and the twist ventral activator region is not ventrally active in the early embryo, demonstrating that repression is dominant over activation. Thus, the default mode of action of the dorsal protein is transcriptional activation. Additional factors may modify dorsal activity to bring about repression. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 1582412 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1672737 # MOTIF: [JP] MA0102.1 Cebpa # MOTIF: [JP] MA0102.2 CEBPA ---------------------------------------------- Nucleic Acids Res. 1991 Jan 11;19(1):131-9. Cell-type specific activity of two glucocorticoid responsive units of rat tyrosine aminotransferase gene is associated with multiple binding sites for C/EBP and a novel liver-specific nuclear factor. Grange T, Roux J, Rigaud G, Pictet R. Institut Jacques Monod du CNRS, Universite Paris, France. The structures of two remote glucocorticoid responsive units (GRUs) that cooperatively interact to promote cell-type specific glucocorticoid induction of rat tyrosine aminotransferase gene expression have been analyzed. DNAase I footprinting and gel mobility shift analyses reveal a complex array of contiguous and overlapping sites for cell type-specific DNA binding proteins. Apart from the glucocorticoid receptor, two liver-specific nuclear factors possess multiple binding sites in each of these GRUs: C/EBP and a newly identified liver-specific factor: HNF5. C/EBP possesses four binding sites in each GRU; a DNA-binding protein with similar binding specificity has been identified in fibroblasts; this protein could be related to AP-3. HNF5 possesses two binding sites in one GRU and four in the other. There are also HNF5 binding sites in numerous regulatory regions of other liver-specific genes. The interaction of HNF5 with DNA gives a characteristic DNAase I footprint with hypersensitive sites in the middle of the recognition sequence. Some of the C/EBP and HNF5 binding sites overlap in a conserved arrangement. Publication Types: Research Support, Non-U.S. Gov't PMID: 1672737 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7592839 # MOTIF: [JP] MA0004.1 Arnt # MOTIF: [JP] MA0006.1 Arnt::Ahr ---------------------------------------------- J Biol Chem. 1995 Nov 3;270(44):26292-302. DNA binding specificities and pairing rules of the Ah receptor, ARNT, and SIM proteins. Swanson HI, Chan WK, Bradfield CA. Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, Chicago, Illinois 60611, USA. The Ah receptor (AHR), the Ah receptor nuclear translocator protein (ARNT), and single-minded protein (SIM) are members of the basic helix-loop-helix-PAS (bHLH-PAS) family of regulatory proteins. In this study, we examine the DNA half-site recognition and pairing rules for these proteins using oligonucleotide selection-amplification and coprecipitation protocols. Oligonucleotide selection-amplification revealed that a variety of bHLH-PAS protein combinations could interact, with each generating a unique DNA binding specificity. To validate the selection-amplification protocol, we demonstrated the preference of the AHR.ARNT complex for the sequence commonly found in dioxin-responsive enhancers in vivo (TNGCGTG). We then demonstrated that the ARNT protein is capable of forming a homodimer with a binding preference for the palindromic E-box sequence, CACGTG. Further examination indicated that ARNT may have a relaxed partner specificity, since it was also capable of forming a heterodimer with SIM and recognizing the sequence GT(G/A)CGTG. Coprecipitation experiments using various PAS proteins and ARNT were consistent with the idea that the ARNT protein has a broad range of interactions among the bHLH-PAS proteins, while the other members appear more restricted in their interactions. Comparison of this in vitro data with sites known to be bound in vivo suggests that the high affinity half-site recognition sequences for the AHR, SIM, and ARNT are T(C/T)GC, GT(G/A)C (5'-half-sites), and GTG (3'-half-sites), respectively. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 7592839 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8062827 # MOTIF: [JP] MA0010.1 br_Z1 # MOTIF: [JP] MA0011.1 br_Z2 # MOTIF: [JP] MA0012.1 br_Z3 # MOTIF: [JP] MA0013.1 br_Z4 ---------------------------------------------- EMBO J. 1994 Aug 1;13(15):3505-16. The Broad-Complex directly controls a tissue-specific response to the steroid hormone ecdysone at the onset of Drosophila metamorphosis. von Kalm L, Crossgrove K, Von Seggern D, Guild GM, Beckendorf SK. Department of Molecular and Cell Biology, University of California, Berkeley 94720. In Drosophila, all of the major metamorphic transitions are regulated by changes in the titer of the steroid hormone ecdysone. Here we examine how a key regulator of metamorphosis and primary ecdysone response gene, the Broad-Complex, transmits the hormonal signal to one of its targets, the Sgs-4 glue gene. We show that Broad-Complex RNAs accumulate in mid third instar larval salivary glands prior to Sgs-4 induction, as expected for the products of a gene that regulates the timing of Sgs-4 activation. The Broad-Complex codes for a family of zinc finger transcriptional regulators. We have identified a number of binding sites for these proteins in sequences known to regulate the timing of Sgs-4 induction, and have used these sites to derive a binding consensus for each protein. Some of these binding sites are required in vivo for Sgs-4 activity. In addition, rbp+, a genetically defined Broad-Complex function that is required for Sgs-4 induction, acts through these Broad-Complex binding sites. Thus, the Broad-Complex directly mediates a temporal and tissue-specific response to ecdysone as larvae become committed to metamorphosis. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S. PMID: 8062827 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8406007 # MOTIF: [JP] MA0014.1 Pax5 ---------------------------------------------- Genes Dev. 1993 Oct;7(10):2048-61. DNA sequence recognition by Pax proteins: bipartite structure of the paired domain and its binding site. Czerny T, Schaffner G, Busslinger M. Research Institute of Molecular Pathology, Vienna, Austria. Previous DNA-binding studies indicated that an intact paired domain is required for interaction of the transcription factor BSAP (Pax-5) with DNA. We have now identified a subset of BSAP recognition sequences that also bind to a truncated BSAP peptide lacking 36 carboxy-terminal amino acids of the paired domain. Sequence comparison of this class of BSAP-binding sites made it possible to unequivocally align all known BSAP-binding sites and to deduce a consensus sequence consisting of two distinct half sites. We propose here a model for the paired domain--DNA interaction in which the paired domain is composed of two subdomains that bind to the two half-sites in adjacent major grooves on the same side of the DNA helix. The existence of these half sites and of the two paired domain subregions was directly demonstrated by methylation interference analysis and by in vitro mutagenesis of both the paired domain and its recognition sequence. Both half-sites contribute to the overall affinity of a given BSAP-binding site according to their match with the consensus sequence. However, none of the naturally occurring BSAP-binding sites completely conform to the consensus sequence. Instead, they contain compensatory base changes in their half-sites that explain the versatile and seemingly degenerate DNA sequence recognition of Pax proteins. Domain swap experiments between BSAP and Pax-1 demonstrated that the sequence specificity of the BSAP paired domain is determined by both its amino- and carboxy-terminal subdomains. Moreover, mutations affecting only one of the two subdomains restricted the sequence specificity of the paired domain. Such mutations have been shown previously to be the cause of mouse developmental mutants (undulated, Splotch, and Small eye) and human syndromes (Waardenburg's syndrome and aniridia) and may thus differentially affect the regulation of target genes by the mutated Pax protein. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8406007 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9973626 # MOTIF: [JP] MA0077.1 SOX9 # MOTIF: [JP] MA0127.1 PEND ---------------------------------------------- Nucleic Acids Res. 1999 Mar 1;27(5):1359-64. The DNA-binding specificity of SOX9 and other SOX proteins. Mertin S, McDowall SG, Harley VR. The Howard Florey Institute of Experimental Physiology and Medicine, University of Melbourne, Parkville, Victoria 3052, Australia. SOX (SRY-related HMG box) proteins are transcription factors that have critical roles in the regulation of numerous developmental processes. They share at least 50% homology in their HMG domains, which bind the DNA element AACAAT. How different SOX proteins achieve specific regulation of target genes is not known. We determined the DNA-binding specificity of SOX9 using a random oligonucleotide selection assay. The optimal SOX9 binding sequence, AGAACAATGG, contained a core DNA-binding element AACAAT, flanked by 5' AG and 3' GG nucleotides. The specific interaction between SOX9 and AGAACAATGG was confirmed by mobility shift assays, DNA competition and dissociation studies. The 5' AG and 3' GG flanking nucleotides enhance binding by SOX9 HMG domain, but not by the HMG domain of another SOX factor, SRY. For SRY, different 5' and 3' flanking nucleotides are preferred. Our studies support the notion that SOX proteins achieve DNA sequence specificity through subtle preferences for flanking nucleotides and that this is likely to be dictated by signature amino acids in their HMG domains. Furthermore, the related HMG domains of SOX9 and Sox17 have similar optimal binding sites that differ from those of SRY and Sox5, suggesting that SOX factors may co-evolve with their DNA targets to achieve specificity. Publication Types: Research Support, Non-U.S. Gov't PMID: 9973626 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8449662 # MOTIF: [JP] MA0061.1 NF-kappaB ---------------------------------------------- Int Rev Cytol. 1993;143:1-62. NF-kappa B and Rel: participants in a multiform transcriptional regulatory system. Grilli M, Chiu JJ, Lenardo MJ. Laboratory of Immunology, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Review PMID: 8449662 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8139574 # MOTIF: [JP] MA0040.1 Foxq1 # MOTIF: [JP] MA0041.1 Foxd3 # MOTIF: [JP] MA0047.1 Foxa2 ---------------------------------------------- Mol Cell Biol. 1994 Apr;14(4):2755-66. The DNA-binding specificity of the hepatocyte nuclear factor 3/forkhead domain is influenced by amino-acid residues adjacent to the recognition helix. Overdier DG, Porcella A, Costa RH. Department of Biochemistry, College of Medicine, University of Illinois at Chicago 60612-7334. Three distinct hepatocyte nuclear factor 3 (HNF-3) proteins (HNF-3 alpha, -3 beta, and -3 gamma) are known to regulate the transcription of liver-specific genes. The HNF-3 proteins bind to DNA as a monomer through a modified helix-turn-helix, known as the winged helix motif, which is also utilized by a number of developmental regulators, including the Drosophila homeotic forkhead (fkh) protein. We have previously described the isolation, from rodent tissue, of an extensive family of tissue-specific HNF-3/fkh homolog (HFH) genes sharing homology in their winged helix motifs. In this report, we have determined the preferred DNA-binding consensus sequence for the HNF-3 beta protein as well as for two divergent family members, HFH-1 and HFH-2. We show that these HNF-3/fkh proteins bind to distinct DNA sites and that the specificity of protein recognition is dependent on subtle nucleotide alterations in the site. The HNF-3, HFH-1, and HFH-2 consensus binding sequences were also used to search DNA regulatory regions to identify potential target genes. Furthermore, an analysis of the DNA-binding properties of a series of HFH-1/HNF-3 beta protein chimeras has allowed us to identify a 20-amino-acid region, located adjacent to the DNA recognition helix, which contributes to DNA-binding specificity. These sequences are not involved in base-specific contacts and include residues which diverge within the HNF-3/fkh family. Replacement of this 20-amino-acid region in HNF-3 beta with corresponding residues from HFH-1 enabled the HNF-3 beta recognition helix to bind only HFH-1-specific DNA-binding sites. We propose a model in which this 20-amino-acid flanking region influences the DNA-binding properties of the recognition helix. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8139574 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7910944 # MOTIF: [JP] MA0070.1 PBX1 ---------------------------------------------- Mol Cell Biol. 1994 Jun;14(6):3938-48. Fusion with E2A converts the Pbx1 homeodomain protein into a constitutive transcriptional activator in human leukemias carrying the t(1;19) translocation. Lu Q, Wright DD, Kamps MP. Department of Chemistry, University of California, San Diego, La Jolla 92093. E2A-PBX1 is a chimeric gene formed by the t(1;19)(q23;p13.3) chromosomal translocation of pediatric pre-B-cell leukemia. The E2A-Pbx1 fusion protein contains sequences encoding the transactivation domain of E2A joined to a majority of the Pbx1 protein, which contains a novel homeodomain. Earlier, we found that expression of E2A-Pbx1 causes malignant transformation of NIH 3T3 fibroblasts and induces myeloid leukemia in mice. Here we demonstrate that the homeodomains encoded by PBX1, as well as by the highly related PBX2 and PBX3 genes, bind the DNA sequence ATCAATCAA. E2A-Pbx1 strongly activates transcription in vivo through this motif, while Pbx1 does not. This finding suggests that E2A-Pbx1 transforms cells by constitutively activating transcription of genes regulated by Pbx1 or by other members of the Pbx protein family. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 7910944 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9420329 # MOTIF: [JP] POL006.1 BREu ---------------------------------------------- Genes Dev. 1998 Jan 1;12(1):34-44. New core promoter element in RNA polymerase II-dependent transcription: sequence-specific DNA binding by transcription factor IIB. Lagrange T, Kapanidis AN, Tang H, Reinberg D, Ebright RH. Howard Hughes Medical Institute and Department of Biochemistry, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA. A sequence element located immediately upstream of the TATA element, and having the consensus sequence 5'-G/C-G/C-G/A-C-G-C-C-3', affects the ability of transcription factor IIB to enter transcription complexes and support transcription initiation. The sequence element is recognized directly by the transcription factor IIB. Recognition involves alpha-helices 4' and 5' of IIB, which comprise a helix-turn-helix DNA-binding motif. These observations establish that transcription initiation involves a fourth core promoter element, the IIB recognition element (BRE), in addition to the TATA element, the initiator element, and the downstream promoter element, and involves a second sequence-specific general transcription factor, IIB, in addition to transcription factor IID. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 9420329 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7737128 # MOTIF: [JP] MA0054.1 myb.Ph3 ---------------------------------------------- EMBO J. 1995 Apr 18;14(8):1773-84. Dual DNA binding specificity of a petal epidermis-specific MYB transcription factor (MYB.Ph3) from Petunia hybrida. Solano R, Nieto C, Avila J, Canas L, Diaz I, Paz-Ares J. Departamento de Biologia Vegetal, Centro de Investigaciones Biologicas-CSIC, Madrid, Spain. The MYB.Ph3 protein recognized two DNA sequences that resemble the two known types of MYB DNA binding site: consensus I (MBSI), aaaAaaC(G/C)-GTTA, and consensus II (MBSII), aaaAGTTAGTTA. Optimal MBSI was recognized by animal c-MYB and not by Am305 from Antirrhinum, whereas MBSII showed the reverse behaviour. Different constraints on MYB.Ph3 binding to the two classes of sequences were demonstrated. DNA binding studies with mutated MBSI and MBSII and hydroxyl radical footprinting analysis, pointed to the N-terminal MYB repeat (R2) as the most involved in determining the dual DNA binding specificity of MYB.Ph3 and supported the idea that binding to MBSI and MBSII does not involve alternative orientations of the two repeats of MYB.Ph3. Minimal promoters containing either MBSI and MBSII were activated to the same extent by MYB.Ph3 in yeast, indicating that both types of binding site can be functionally equivalent. MYB.Ph3 binding sites are present in the promoter of flavonoid biosynthetic genes, such as the Petunia chsJ gene, which was transcriptionally activated by MYB.Ph3 in tobacco protoplasts. MYB.Ph3 was immunolocalized in the epidermal cell layer of petals, where flavonoid biosynthetic genes are actively expressed. This strongly suggests a role for MYB.Ph3 in the regulation of flavonoid biosynthesis. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 7737128 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 14502648 # MOTIF: [JP] MA0133.1 BRCA1 ---------------------------------------------- Mol Carcinog. 2003 Oct;38(2):85-96. Novel consensus DNA-binding sequence for BRCA1 protein complexes. Cable PL, Wilson CA, Calzone FJ, Rauscher FJ 3rd, Scully R, Livingston DM, Li L, Blackwell CB, Futreal PA, Afshari CA. Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, USA. Increasing evidence continues to emerge supporting the early hypothesis that BRCA1 might be involved in transcriptional processes. BRCA1 physically associates with more than 15 different proteins involved in transcription and is paradoxically involved in both transcriptional activation and repression. However, the underlying mechanism by which BRCA1 affects the gene expression of various genes remains speculative. In this study, we provide evidence that BRCA1 protein complexes interact with specific DNA sequences. We provide data showing that the upstream stimulatory factor 2 (USF2) physically associates with BRCA1 and is a component of this DNA-binding complex. Interestingly, these DNA-binding complexes are downregulated in breast cancer cell lines containing wild-type BRCA1, providing a critical link between modulations of BRCA1 function in sporadic breast cancers that do not involve germline BRCA1 mutations. The functional specificity of BRCA1 tumor suppression for breast and ovarian tissues is supported by our experiments, which demonstrate that BRCA1 DNA-binding complexes are modulated by serum and estrogen. Finally, functional analysis indicates that missense mutations in BRCA1 that lead to subsequent cancer susceptibility may result in improper gene activation. In summary, these findings establish a role for endogenous BRCA1 protein complexes in transcription via a defined DNA-binding sequence and indicate that one function of BRCA1 is to co-regulate the expression of genes involved in various cellular processes. Published 2003 Wiley-Liss, Inc. PMID: 14502648 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7957066 # MOTIF: [JP] MA0030.1 FOXF2 # MOTIF: [JP] MA0031.1 FOXD1 # MOTIF: [JP] MA0032.1 FOXC1 # MOTIF: [JP] MA0033.1 FOXL1 ---------------------------------------------- EMBO J. 1994 Oct 17;13(20):5002-12. Cloning and characterization of seven human forkhead proteins: binding site specificity and DNA bending. Pierrou S, Hellqvist M, Samuelsson L, Enerback S, Carlsson P. Department of Molecular Biology, Goteborg University, Sweden. The forkhead domain is a monomeric DNA binding motif that defines a rapidly growing family of eukaryotic transcriptional regulators. Genetic and biochemical data suggest a central role in embryonic development for genes encoding forkhead proteins. We have used PCR and low stringency hybridization to isolate clones from human cDNA and genomic libraries that represent seven novel forkhead genes, freac-1 to freac-7. The spatial patterns of expression for the seven freac genes range from specific for a single tissue to nearly ubiquitous. The DNA binding specificities of four of the FREAC proteins were determined by selection of binding sites from random sequence oligonucleotides. The binding sites for all four FREAC proteins share a core sequence, RTAAAYA, but differ in the positions flanking the core. Domain swaps between two FREAC proteins identified two subregions within the forkhead domain as responsible for creating differences in DNA binding specificity. Applying a circular permutation assay, we show that binding of FREAC proteins to their cognate sites results in bending of the DNA at an angle of 80-90 degrees. Publication Types: Research Support, Non-U.S. Gov't PMID: 7957066 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8524663 # MOTIF: [JP] MA0076.1 ELK4 ---------------------------------------------- Nucleic Acids Res. 1995 Nov 25;23(22):4698-706. The ETS-domain transcription factors Elk-1 and SAP-1 exhibit differential DNA binding specificities. Shore P, Sharrocks AD. Department of Biochemistry and Genetics, Medical School, University of Newcastle upon Tyne, UK. The ETS DNA-binding domain is conserved amongst many eukaryotic transcription factors. ETS-domains bind differentially to specific DNA sites containing a central GGA trinucleotide motif. The nucleotides flanking this motif define the binding specificity of individual proteins. In this study we have investigated binding specificity of the ETS-domains from two members of the ternary complex factor (TCF) subfamily, Elk-1 and SAP-1. The ETS DNA-binding domains of Elk-1 (Elk-93) and SAP-1 (SAP-92) select similar sites from random pools of double stranded oligonucleotides based on the consensus sequence ACCGGAAGTR. However, SAP-92 shows a more relaxed binding site selectivity and binds efficiently to a greater spectrum of sites than does Elk-93. This more relaxed DNA binding site selectivity is most pronounced in nucleotides located on the 3' side of the GGA motif. This differential DNA-binding specificity is also exhibited by longer TCF derivatives and, indeed by the full-length proteins. Our results suggest that the range of potential in vivo target sites for SAP-1 is likely to be greater than for Elk-1. We discuss our results in relation to other similar studies carried out with more divergent ETS-domains. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8524663 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 18585360 # MOTIF: [JP] MA0094.2 Ubx # MOTIF: [JP] MA0180.1 Vsx2 # MOTIF: [JP] MA0183.1 CG7056 # MOTIF: [JP] MA0187.1 Dll # MOTIF: [JP] MA0197.1 Oct # MOTIF: [JP] MA0201.1 Ptx1 # MOTIF: [JP] MA0208.1 al # MOTIF: [JP] MA0222.1 exd # MOTIF: [JP] MA0224.1 exex # MOTIF: [JP] MA0229.1 inv # MOTIF: [JP] MA0247.1 tin # MOTIF: [JP] MA0253.1 vnd ---------------------------------------------- Cell. 2008 Jun 27;133(7):1277-89. Comment in: Cell. 2008 Jun 27;133(7):1133-5. Analysis of homeodomain specificities allows the family-wide prediction of preferred recognition sites. Noyes MB, Christensen RG, Wakabayashi A, Stormo GD, Brodsky MH, Wolfe SA. Program in Gene Function and Expression, University of Massachusetts Medical School, Worcester, MA 01605, USA. We describe the comprehensive characterization of homeodomain DNA-binding specificities from a metazoan genome. The analysis of all 84 independent homeodomains from D. melanogaster reveals the breadth of DNA sequences that can be specified by this recognition motif. The majority of these factors can be organized into 11 different specificity groups, where the preferred recognition sequence between these groups can differ at up to four of the six core recognition positions. Analysis of the recognition motifs within these groups led to a catalog of common specificity determinants that may cooperate or compete to define the binding site preference. With these recognition principles, a homeodomain can be reengineered to create factors where its specificity is altered at the majority of recognition positions. This resource also allows prediction of homeodomain specificities from other organisms, which is demonstrated by the prediction and analysis of human homeodomain specificities. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 18585360 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 10497269 # MOTIF: [JP] MA0003.1 TFAP2A ---------------------------------------------- Nucleic Acids Res. 1999 Oct 15;27(20):4040-9. AP2alpha and AP2gamma: a comparison of binding site specificity and trans-activation of the estrogen receptor promoter and single site promoter constructs. McPherson LA, Weigel RJ. Department of Surgery, Stanford University, MSLS, Room P214, Stanford, CA 94305-5408, USA. The AP2 transcription factors exhibit a high degree of homology in the DNA binding and dimerization domains. In this study, we methodically compared the binding specificity of AP2alpha and AP2gamma using PCR-assisted binding site selection and competitive gel shift assay and determined that the consensus binding site for both factors is(G)/(C)CCNN(A/)C(/G)(G)/(A)G(G/)C(/T.)The use of single site promoter constructs with either a high or low affinity site demonstrated a direct relationship between site affinity and transcriptional activation. Overexpression of AP2alpha and AP2gamma resulted in the activation of a low affinity binding site construct to levels comparable to those seen with a high affinity site construct at lower amounts of protein expression. Both AP2alpha and AP2gamma were able to trans-activate the cloned human estrogen receptor alpha promoter in ER-negative MDA-MB-231 cells through high affinity AP2 sites in the untranslated leader sequence. This provides a functional mechanism to explain the correlation between AP2 activity and estrogen receptor expression in breast cancer. Since there is overexpression of AP2 factors in breast cancer compared to normal breast epithelium, our results suggest that increased factor expression may activate a set of target genes containing lower affinity binding sites that would normally not be expressed in normal breast epithelium. Publication Types: Comparative Study Research Support, U.S. Gov't, P.H.S. PMID: 10497269 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1588974 # MOTIF: [JP] MA0106.1 TP53 ---------------------------------------------- Mol Cell Biol. 1992 Jun;12(6):2866-71. A transcriptionally active DNA-binding site for human p53 protein complexes. Funk WD, Pak DT, Karas RH, Wright WE, Shay JW. Department of Cell Biology and Neuroscience, University of Texas Southwestern Medical Center, Dallas 75235-9039. Recent studies have demonstrated transcriptional activation domains within the tumor suppressor protein p53, while others have described specific DNA-binding sites for p53, implying that the protein may act as a transcriptional regulatory factor. We have used a reiterative selection procedure (CASTing: cyclic amplification and selection of targets) to identify new specific binding sites for p53, using nuclear extracts from normal human fibroblasts as the source of p53 protein. The preferred consensus is the palindrome GGACATGCCCGGGCATGTCC. In vitro-translated p53 binds to this sequence only when mixed with nuclear extracts, suggesting that p53 may bind DNA after posttranslational modification or as a complex with other protein partners. When placed upstream of a reporter construct, this sequence promotes p53-dependent transcription in transient transfection assays. Publication Types: In Vitro Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 1588974 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8065305 # MOTIF: [JP] MA0103.1 ZEB1 ---------------------------------------------- Mol Cell Biol. 1994 Sep;14(9):5692-700. The delta-crystallin enhancer-binding protein delta EF1 is a repressor of E2-box-mediated gene activation. Sekido R, Murai K, Funahashi J, Kamachi Y, Fujisawa-Sehara A, Nabeshima Y, Kondoh H. Institute for Molecular and Cellular Biology, Osaka University, Japan. The repressor delta EF1 was discovered by its action on the DC5 fragment of the lens-specific delta 1-crystallin enhancer. C-proximal zinc fingers of delta EF1 were found responsible for binding to the DC5 fragment and had specificity to CACCT as revealed by selection of high-affinity binding sequences from a random oligonucleotide pool. CACCT is present not only in DC5 but also in the E2 box (CACCTG) elements which are the binding sites of various basic helix-loop-helix activators and also the target of an unidentified repressor, raising the possibility that delta EF1 accounts for the E2 box repressor activity. delta EF1 competed with E47 for binding to an E2 box sequence in vitro. In lymphoid cells, endogenous delta EF1 activity as a repressor was detectable, and exogenous delta EF1 repressed immunoglobulin kappa enhancer by binding to the kappa E2 site. Moreover, delta EF1 repressed MyoD-dependent activation of the muscle creatine kinase enhancer and MyoD-induced myogenesis of 10T1/2 cells. Thus, delta EF1 counteracts basic helix-loop-helix activators through binding site competition and fulfills the conditions of the E2 box repressor. In embryonic tissues, the most prominent site of delta EF1 expression is the myotome. Myotomal expression as well as the above results argues for a significant contribution of delta EF1 in regulation of embryonic myogenesis through the modulation of the actions of MyoD family proteins. Publication Types: Comparative Study Research Support, Non-U.S. Gov't PMID: 8065305 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 16041365 # MOTIF: [JP] MA0162.1 Egr1 # MOTIF: [JP] MA0163.1 PLAG1 # MOTIF: [JP] MA0239.1 prd # MOTIF: [JP] MA0242.1 run::Bgb ---------------------------------------------- Nat Biotechnol. 2005 Aug;23(8):988-94. Epub 2005 Jul 24. A bacterial one-hybrid system for determining the DNA-binding specificity of transcription factors. Meng X, Brodsky MH, Wolfe SA. Program in Gene Function and Expression, University of Massachusetts Medical School, 364 Plantation St., Worcester, Massachusetts 01605, USA. The DNA-binding specificities of transcription factors can be used to computationally predict cis-regulatory modules (CRMs) that regulate gene expression. However, the absence of specificity data for the majority of transcription factors limits the widespread implementation of this approach. We have developed a bacterial one-hybrid system that provides a simple and rapid method to determine the DNA-binding specificity of a transcription factor. Using this technology, we successfully determined the DNA-binding specificity of seven previously characterized transcription factors and one novel transcription factor, the Drosophila melanogaster factor Odd-skipped. Regulatory targets of Odd-skipped were successfully predicted using this information, demonstrating that the data produced by the bacterial one-hybrid system are relevant to in vivo function. Publication Types: Evaluation Studies Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 16041365 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19443739 # MOTIF: [JP] PB0001.1 Arid3a_1 # MOTIF: [JP] PB0002.1 Arid5a_1 # MOTIF: [JP] PB0003.1 Ascl2_1 # MOTIF: [JP] PB0004.1 Atf1_1 # MOTIF: [JP] PB0005.1 Bbx_1 # MOTIF: [JP] PB0006.1 Bcl6b_1 # MOTIF: [JP] PB0007.1 Bhlhb2_1 # MOTIF: [JP] PB0008.1 E2F2_1 # MOTIF: [JP] PB0009.1 E2F3_1 # MOTIF: [JP] PB0010.1 Egr1_1 # MOTIF: [JP] PB0011.1 Ehf_1 # MOTIF: [JP] PB0012.1 Elf3_1 # MOTIF: [JP] PB0013.1 Eomes_1 # MOTIF: [JP] PB0014.1 Esrra_1 # MOTIF: [JP] PB0015.1 Foxa2_1 # MOTIF: [JP] PB0016.1 Foxj1_1 # MOTIF: [JP] PB0017.1 Foxj3_1 # MOTIF: [JP] PB0018.1 Foxk1_1 # MOTIF: [JP] PB0019.1 Foxl1_1 # MOTIF: [JP] PB0020.1 Gabpa_1 # MOTIF: [JP] PB0021.1 Gata3_1 # MOTIF: [JP] PB0022.1 Gata5_1 # MOTIF: [JP] PB0023.1 Gata6_1 # MOTIF: [JP] PB0024.1 Gcm1_1 # MOTIF: [JP] PB0025.1 Glis2_1 # MOTIF: [JP] PB0026.1 Gm397_1 # MOTIF: [JP] PB0027.1 Gmeb1_1 # MOTIF: [JP] PB0028.1 Hbp1_1 # MOTIF: [JP] PB0029.1 Hic1_1 # MOTIF: [JP] PB0030.1 Hnf4a_1 # MOTIF: [JP] PB0031.1 Hoxa3_1 # MOTIF: [JP] PB0032.1 IRC900814_1 # MOTIF: [JP] PB0033.1 Irf3_1 # MOTIF: [JP] PB0034.1 Irf4_1 # MOTIF: [JP] PB0035.1 Irf5_1 # MOTIF: [JP] PB0036.1 Irf6_1 # MOTIF: [JP] PB0037.1 Isgf3g_1 # MOTIF: [JP] PB0038.1 Jundm2_1 # MOTIF: [JP] PB0039.1 Klf7_1 # MOTIF: [JP] PB0040.1 Lef1_1 # MOTIF: [JP] PB0041.1 Mafb_1 # MOTIF: [JP] PB0042.1 Mafk_1 # MOTIF: [JP] PB0043.1 Max_1 # MOTIF: [JP] PB0044.1 Mtf1_1 # MOTIF: [JP] PB0045.1 Myb_1 # MOTIF: [JP] PB0046.1 Mybl1_1 # MOTIF: [JP] PB0047.1 Myf6_1 # MOTIF: [JP] PB0048.1 Nkx3-1_1 # MOTIF: [JP] PB0049.1 Nr2f2_1 # MOTIF: [JP] PB0050.1 Osr1_1 # MOTIF: [JP] PB0051.1 Osr2_1 # MOTIF: [JP] PB0052.1 Plagl1_1 # MOTIF: [JP] PB0053.1 Rara_1 # MOTIF: [JP] PB0054.1 Rfx3_1 # MOTIF: [JP] PB0055.1 Rfx4_1 # MOTIF: [JP] PB0056.1 Rfxdc2_1 # MOTIF: [JP] PB0057.1 Rxra_1 # MOTIF: [JP] PB0058.1 Sfpi1_1 # MOTIF: [JP] PB0059.1 Six6_1 # MOTIF: [JP] PB0060.1 Smad3_1 # MOTIF: [JP] PB0061.1 Sox11_1 # MOTIF: [JP] PB0062.1 Sox12_1 # MOTIF: [JP] PB0063.1 Sox13_1 # MOTIF: [JP] PB0064.1 Sox14_1 # MOTIF: [JP] PB0065.1 Sox15_1 # MOTIF: [JP] PB0066.1 Sox17_1 # MOTIF: [JP] PB0067.1 Sox18_1 # MOTIF: [JP] PB0068.1 Sox1_1 # MOTIF: [JP] PB0069.1 Sox21_1 # MOTIF: [JP] PB0070.1 Sox30_1 # MOTIF: [JP] PB0071.1 Sox4_1 # MOTIF: [JP] PB0072.1 Sox5_1 # MOTIF: [JP] PB0073.1 Sox7_1 # MOTIF: [JP] PB0074.1 Sox8_1 # MOTIF: [JP] PB0075.1 Sp100_1 # MOTIF: [JP] PB0076.1 Sp4_1 # MOTIF: [JP] PB0077.1 Spdef_1 # MOTIF: [JP] PB0078.1 Srf_1 # MOTIF: [JP] PB0079.1 Sry_1 # MOTIF: [JP] PB0080.1 Tbp_1 # MOTIF: [JP] PB0081.1 Tcf1_1 # MOTIF: [JP] PB0082.1 Tcf3_1 # MOTIF: [JP] PB0083.1 Tcf7_1 # MOTIF: [JP] PB0084.1 Tcf7l2_1 # MOTIF: [JP] PB0085.1 Tcfap2a_1 # MOTIF: [JP] PB0086.1 Tcfap2b_1 # MOTIF: [JP] PB0087.1 Tcfap2c_1 # MOTIF: [JP] PB0088.1 Tcfap2e_1 # MOTIF: [JP] PB0089.1 Tcfe2a_1 # MOTIF: [JP] PB0090.1 Zbtb12_1 # MOTIF: [JP] PB0091.1 Zbtb3_1 # MOTIF: [JP] PB0092.1 Zbtb7b_1 # MOTIF: [JP] PB0093.1 Zfp105_1 # MOTIF: [JP] PB0094.1 Zfp128_1 # MOTIF: [JP] PB0095.1 Zfp161_1 # MOTIF: [JP] PB0096.1 Zfp187_1 # MOTIF: [JP] PB0097.1 Zfp281_1 # MOTIF: [JP] PB0098.1 Zfp410_1 # MOTIF: [JP] PB0099.1 Zfp691_1 # MOTIF: [JP] PB0100.1 Zfp740_1 # MOTIF: [JP] PB0101.1 Zic1_1 # MOTIF: [JP] PB0102.1 Zic2_1 # MOTIF: [JP] PB0103.1 Zic3_1 # MOTIF: [JP] PB0104.1 Zscan4_1 # MOTIF: [JP] PB0105.1 Arid3a_2 # MOTIF: [JP] PB0106.1 Arid5a_2 # MOTIF: [JP] PB0107.1 Ascl2_2 # MOTIF: [JP] PB0108.1 Atf1_2 # MOTIF: [JP] PB0109.1 Bbx_2 # MOTIF: [JP] PB0110.1 Bcl6b_2 # MOTIF: [JP] PB0111.1 Bhlhb2_2 # MOTIF: [JP] PB0112.1 E2F2_2 # MOTIF: [JP] PB0113.1 E2F3_2 # MOTIF: [JP] PB0114.1 Egr1_2 # MOTIF: [JP] PB0115.1 Ehf_2 # MOTIF: [JP] PB0116.1 Elf3_2 # MOTIF: [JP] PB0117.1 Eomes_2 # MOTIF: [JP] PB0118.1 Esrra_2 # MOTIF: [JP] PB0119.1 Foxa2_2 # MOTIF: [JP] PB0120.1 Foxj1_2 # MOTIF: [JP] PB0121.1 Foxj3_2 # MOTIF: [JP] PB0122.1 Foxk1_2 # MOTIF: [JP] PB0123.1 Foxl1_2 # MOTIF: [JP] PB0124.1 Gabpa_2 # MOTIF: [JP] PB0125.1 Gata3_2 # MOTIF: [JP] PB0126.1 Gata5_2 # MOTIF: [JP] PB0127.1 Gata6_2 # MOTIF: [JP] PB0128.1 Gcm1_2 # MOTIF: [JP] PB0129.1 Glis2_2 # MOTIF: [JP] PB0130.1 Gm397_2 # MOTIF: [JP] PB0131.1 Gmeb1_2 # MOTIF: [JP] PB0132.1 Hbp1_2 # MOTIF: [JP] PB0133.1 Hic1_2 # MOTIF: [JP] PB0134.1 Hnf4a_2 # MOTIF: [JP] PB0135.1 Hoxa3_2 # MOTIF: [JP] PB0136.1 IRC900814_2 # MOTIF: [JP] PB0137.1 Irf3_2 # MOTIF: [JP] PB0138.1 Irf4_2 # MOTIF: [JP] PB0139.1 Irf5_2 # MOTIF: [JP] PB0140.1 Irf6_2 # MOTIF: [JP] PB0141.1 Isgf3g_2 # MOTIF: [JP] PB0142.1 Jundm2_2 # MOTIF: [JP] PB0143.1 Klf7_2 # MOTIF: [JP] PB0144.1 Lef1_2 # MOTIF: [JP] PB0145.1 Mafb_2 # MOTIF: [JP] PB0146.1 Mafk_2 # MOTIF: [JP] PB0147.1 Max_2 # MOTIF: [JP] PB0148.1 Mtf1_2 # MOTIF: [JP] PB0149.1 Myb_2 # MOTIF: [JP] PB0150.1 Mybl1_2 # MOTIF: [JP] PB0151.1 Myf6_2 # MOTIF: [JP] PB0152.1 Nkx3-1_2 # MOTIF: [JP] PB0153.1 Nr2f2_2 # MOTIF: [JP] PB0154.1 Osr1_2 # MOTIF: [JP] PB0155.1 Osr2_2 # MOTIF: [JP] PB0156.1 Plagl1_2 # MOTIF: [JP] PB0157.1 Rara_2 # MOTIF: [JP] PB0158.1 Rfx3_2 # MOTIF: [JP] PB0159.1 Rfx4_2 # MOTIF: [JP] PB0160.1 Rfxdc2_2 # MOTIF: [JP] PB0161.1 Rxra_2 # MOTIF: [JP] PB0162.1 Sfpi1_2 # MOTIF: [JP] PB0163.1 Six6_2 # MOTIF: [JP] PB0164.1 Smad3_2 # MOTIF: [JP] PB0165.1 Sox11_2 # MOTIF: [JP] PB0166.1 Sox12_2 # MOTIF: [JP] PB0167.1 Sox13_2 # MOTIF: [JP] PB0168.1 Sox14_2 # MOTIF: [JP] PB0169.1 Sox15_2 # MOTIF: [JP] PB0170.1 Sox17_2 # MOTIF: [JP] PB0171.1 Sox18_2 # MOTIF: [JP] PB0172.1 Sox1_2 # MOTIF: [JP] PB0173.1 Sox21_2 # MOTIF: [JP] PB0174.1 Sox30_2 # MOTIF: [JP] PB0175.1 Sox4_2 # MOTIF: [JP] PB0176.1 Sox5_2 # MOTIF: [JP] PB0177.1 Sox7_2 # MOTIF: [JP] PB0178.1 Sox8_2 # MOTIF: [JP] PB0179.1 Sp100_2 # MOTIF: [JP] PB0180.1 Sp4_2 # MOTIF: [JP] PB0181.1 Spdef_2 # MOTIF: [JP] PB0182.1 Srf_2 # MOTIF: [JP] PB0183.1 Sry_2 # MOTIF: [JP] PB0184.1 Tbp_2 # MOTIF: [JP] PB0185.1 Tcf1_2 # MOTIF: [JP] PB0186.1 Tcf3_2 # MOTIF: [JP] PB0187.1 Tcf7_2 # MOTIF: [JP] PB0188.1 Tcf7l2_2 # MOTIF: [JP] PB0189.1 Tcfap2a_2 # MOTIF: [JP] PB0190.1 Tcfap2b_2 # MOTIF: [JP] PB0191.1 Tcfap2c_2 # MOTIF: [JP] PB0192.1 Tcfap2e_2 # MOTIF: [JP] PB0193.1 Tcfe2a_2 # MOTIF: [JP] PB0194.1 Zbtb12_2 # MOTIF: [JP] PB0195.1 Zbtb3_2 # MOTIF: [JP] PB0196.1 Zbtb7b_2 # MOTIF: [JP] PB0197.1 Zfp105_2 # MOTIF: [JP] PB0198.1 Zfp128_2 # MOTIF: [JP] PB0199.1 Zfp161_2 # MOTIF: [JP] PB0200.1 Zfp187_2 # MOTIF: [JP] PB0201.1 Zfp281_2 # MOTIF: [JP] PB0202.1 Zfp410_2 # MOTIF: [JP] PB0203.1 Zfp691_2 # MOTIF: [JP] PB0204.1 Zfp740_2 # MOTIF: [JP] PB0205.1 Zic1_2 # MOTIF: [JP] PB0206.1 Zic2_2 # MOTIF: [JP] PB0207.1 Zic3_2 # MOTIF: [JP] PB0208.1 Zscan4_2 ---------------------------------------------- Science. 2009 Jun 26;324(5935):1720-3. Epub 2009 May 14. Diversity and complexity in DNA recognition by transcription factors. Badis G, Berger MF, Philippakis AA, Talukder S, Gehrke AR, Jaeger SA, Chan ET, Metzler G, Vedenko A, Chen X, Kuznetsov H, Wang CF, Coburn D, Newburger DE, Morris Q, Hughes TR, Bulyk ML. Banting and Best Department of Medical Research, University of Toronto, Toronto, ON M5S 3E1, Canada. Sequence preferences of DNA binding proteins are a primary mechanism by which cells interpret the genome. Despite the central importance of these proteins in physiology, development, and evolution, comprehensive DNA binding specificities have been determined experimentally for only a few proteins. Here, we used microarrays containing all 10-base pair sequences to examine the binding specificities of 104 distinct mouse DNA binding proteins representing 22 structural classes. Our results reveal a complex landscape of binding, with virtually every protein analyzed possessing unique preferences. Roughly half of the proteins each recognized multiple distinctly different sequence motifs, challenging our molecular understanding of how proteins interact with their DNA binding sites. This complexity in DNA recognition may be important in gene regulation and in the evolution of transcriptional regulatory networks. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 19443739 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9571041 # MOTIF: [JP] MA0055.1 Myf # MOTIF: [JP] MA0090.1 TEAD1 ---------------------------------------------- J Mol Biol. 1998 Apr 24;278(1):167-81. Identification of regulatory regions which confer muscle-specific gene expression. Wasserman WW, Fickett JW. Bioinformatics Research Group, SmithKline Beecham Pharmaceuticals, 709 Swedeland Road, King of Prussia, PA 19406, USA. For many newly sequenced genes, sequence analysis of the putative protein yields no clue on function. It would be beneficial to be able to identify in the genome the regulatory regions that confer temporal and spatial expression patterns for the uncharacterized genes. Additionally, it would be advantageous to identify regulatory regions within genes of known expression pattern without performing the costly and time consuming laboratory studies now required. To achieve these goals, the wealth of case studies performed over the past 15 years will have to be collected into predictive models of expression. Extensive studies of genes expressed in skeletal muscle have identified specific transcription factors which bind to regulatory elements to control gene expression. However, potential binding sites for these factors occur with sufficient frequency that it is rare for a gene to be found without one. Analysis of experimentally determined muscle regulatory sequences indicates that muscle expression requires multiple elements in close proximity. A model is generated with predictive capability for identifying these muscle-specific regulatory modules. Phylogenetic footprinting, the identification of sequences conserved between distantly related species, complements the statistical predictions. Through the use of logistic regression analysis, the model promises to be easily modified to take advantage of the elucidation of additional factors, cooperation rules, and spacing constraints. Copyright 1998 Academic Press Limited. Publication Types: Research Support, U.S. Gov't, P.H.S. PMID: 9571041 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 19160518 # MOTIF: [JP] MA0062.2 GABPA ---------------------------------------------- Nat Methods. 2008 Sep;5(9):829-34. Genome-wide analysis of transcription factor binding sites based on ChIP-Seq data. Valouev A, Johnson DS, Sundquist A, Medina C, Anton E, Batzoglou S, Myers RM, Sidow A. Department of Pathology, Stanford University Medical Center, 300 Pasteur Drive, Stanford, California 94305, USA. Molecular interactions between protein complexes and DNA mediate essential gene-regulatory functions. Uncovering such interactions by chromatin immunoprecipitation coupled with massively parallel sequencing (ChIP-Seq) has recently become the focus of intense interest. We here introduce quantitative enrichment of sequence tags (QuEST), a powerful statistical framework based on the kernel density estimation approach, which uses ChIP-Seq data to determine positions where protein complexes contact DNA. Using QuEST, we discovered several thousand binding sites for the human transcription factors SRF, GABP and NRSF at an average resolution of about 20 base pairs. MEME motif-discovery tool-based analyses of the QuEST-identified sequences revealed DNA binding by cofactors of SRF, providing evidence that cofactor binding specificity can be obtained from ChIP-Seq data. By combining QuEST analyses with Gene Ontology (GO) annotations and expression data, we illustrate how general functions of transcription factors can be inferred. Publication Types: Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMID: 19160518 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 7926749 # MOTIF: [JP] MA0071.1 RORA_1 # MOTIF: [JP] MA0072.1 RORA_2 ---------------------------------------------- Genes Dev. 1994 Mar 1;8(5):538-53. Isoform-specific amino-terminal domains dictate DNA-binding properties of ROR alpha, a novel family of orphan hormone nuclear receptors. Giguere V, Tini M, Flock G, Ong E, Evans RM, Otulakowski G. Division of Endocrinology, Hospital for Sick Children, Toronto, Ontario, Canada. Three isoforms of a novel member of the steroid hormone nuclear receptor superfamily related to the retinoic acid receptors have been identified. The three isoforms, referred to as ROR alpha 1, ROR alpha 2, and ROR alpha 3, share common DNA- and putative ligand-binding domains but are characterized by distinct amino-terminal domains generated by alternative RNA processing. An exon encoding a functionally important subregion of the amino-terminal domain of the ROR alpha 2 isoform resides on the opposite strand of a cytochrome c-processed pseudogene. Binding site selection using in vitro-synthesized proteins reveals that the ROR alpha 1 and ROR alpha 2 isoforms bind DNA as monomers to hormone response elements composed of a 6-bp AT-rich sequence preceding a half-site core motif PuGGTCA (RORE). However, ROR alpha 1 and ROR alpha 2 display different binding specificities: ROR alpha 1 binds to and constitutively activates transcription from a large subset of ROREs, whereas ROR alpha 2 recognizes ROREs with strict specificity and displays weaker transcriptional activity. The differential DNA-binding activity of each isoform maps to their respective amino-terminal domains. Whereas truncation of the amino-terminal domain diminishes the ability of ROR alpha 1 to bind DNA, a similar deletion relaxes ROR alpha 2-binding specificity to that displayed by ROR alpha 1. Remarkably, transfer of the entire amino-terminal region of ROR alpha 1 or amino-terminal deletion of ROR alpha 2 confers RORE-binding specificities to heterologous receptors. These results demonstrate that the amino-terminal domain and the zinc finger region work in concert to confer high affinity and specific DNA-binding properties to the ROR isoforms and suggest a novel strategy to control DNA-binding activity of nuclear receptors. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 7926749 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15555547 # MOTIF: [JP] MA0130.1 ZNF354C ---------------------------------------------- Biochem Biophys Res Commun. 2004 Dec 24;325(4):1145-52. Cloning and characterization of a novel human zinc finger gene, hKid3, from a C2H2-ZNF enriched human embryonic cDNA library. Gao L, Sun C, Qiu HL, Liu H, Shao HJ, Wang J, Li WX. Ministry of Education Key Laboratory of Virology, College of Life Science, Wuhan University, Wuhan, Hubei 430072, PR China. liwxlab@whu.edu.cn To investigate the zinc finger genes involved in human embryonic development, we constructed a C(2)H(2)-ZNF enriched human embryonic cDNA library, from which a novel human gene named hKid3 was identified. The hKid3 cDNA encodes a 554 amino acid protein with an amino-terminal KRAB domain and 11 carboxyl-terminal C(2)H(2) zinc finger motifs. Northern blot analysis indicates that two hKid3 transcripts of 6 and 8.5kb express in human fetal brain and kidney. The 6kb transcript can also be detected in human adult brain, heart, and skeletal muscle while the 8.5kb transcript appears to be embryo-specific. GFP-fused hKid3 protein is localized to nuclei and the ZF domain is necessary and sufficient for nuclear localization. To explore the DNA-binding specificity of hKid3, an oligonucleotide library was selected by GST fusion protein of hKid3 ZF domain, and the consensus core sequence 5'-CCAC-3' was evaluated by competitive electrophoretic mobility shift assay. Moreover, The KRAB domain of hKid3 exhibits transcription repressor activity when tested in GAL4 fusion protein assay. These results indicate that hKid3 may function as a transcription repressor with regulated expression pattern during human development of brain and kidney. Publication Types: Comparative Study Evaluation Studies Research Support, Non-U.S. Gov't PMID: 15555547 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1861984 # MOTIF: [JP] MA0100.1 Myb ---------------------------------------------- Nucleic Acids Res. 1991 Jul 25;19(14):3913-9. Nucleotide preferences in sequence-specific recognition of DNA by c-myb protein. Howe KM, Watson RJ. Imperial Cancer Research Fund, Lincoln's Inn Fields, London, UK. Using a binding site selection procedure, we have found that sequence-specific DNA-binding by the mouse c-myb protein involves recognition of nucleotides outside of the previously identified hexanucleotide motif. Oligonucleotides containing a random nucleotide core were immunoprecipitated in association with c-Myb, amplified by the Polymerase Chain Reaction and cloned in plasmids prior to sequencing. By alignment of sequences it was apparent that additional preferences existed at each of three bases immediately 5' of the hexanucleotide consensus, allowing an extension of the preferred binding site to YGRCVGTTR. The contributions of these 5' nucleotides to binding affinity was established in bandshift analyses with oligonucleotides containing single base substitutions; in particular, it was found that replacement of the preferred guanine at position -2 with any other base greatly reduced c-Myb binding. We found that the protein encoded by the related B-myb gene bound the preferred c-Myb site with similar affinity; however, B-Myb and c-Myb showed distinct preferences for the identity of the nucleotide at position -1 relative to the hexanucleotide consensus. This study demonstrates that the c-Myb DNA-binding site is more extensive than recognised hitherto and points to similar but distinct nucleotide preferences in recognition of DNA by related Myb proteins. PMID: 1861984 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 1620116 # MOTIF: [JP] MA0025.1 NFIL3 ---------------------------------------------- Mol Cell Biol. 1992 Jul;12(7):3070-7. Transcriptional repression by a novel member of the bZIP family of transcription factors. Cowell IG, Skinner A, Hurst HC. Gene Transcription Laboratory, Imperial Cancer Research Fund, Hammersmith Hospital, London, England. We describe here a novel member of the bZIP family of DNA-binding proteins, designated E4BP4, that displays an unusual DNA-binding specificity which overlaps that of the activating transcription factor family of factors. When expressed in a transient transfection assay with a suitable reporter plasmid, E4BP4 strongly repressed transcription in a DNA-binding-site-dependent manner. Examination of a series of deletion mutants revealed that sequences responsible for the repressing potential of E4BP4 lie within the carboxyl-terminal region of the protein. No similarity was found between this region and the repressing domains of other known eukaryotic transcriptional repressors. PMID: 1620116 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 15231738 # MOTIF: [JP] POL001.1 MTE ---------------------------------------------- Genes Dev. 2004 Jul 1;18(13):1606-17. The MTE, a new core promoter element for transcription by RNA polymerase II. Lim CY, Santoso B, Boulay T, Dong E, Ohler U, Kadonaga JT. Section of Molecular Biology, University of California at San Diego, La Jolla, California 92093, USA. The core promoter is the ultimate target of the vast network of regulatory factors that contribute to the initiation of transcription by RNA polymerase II. Here we describe the MTE (motif ten element), a new core promoter element that appears to be conserved from Drosophila to humans. The MTE promotes transcription by RNA polymerase II when it is located precisely at positions +18 to +27 relative to A(+1) in the initiator (Inr) element. MTE sequences from +18 to +22 relative to A(+1) are important for basal transcription, and a region from +18 to +27 is sufficient to confer MTE activity to heterologous core promoters. The MTE requires the Inr, but functions independently of the TATA-box and DPE. Notably, the loss of transcriptional activity upon mutation of a TATA-box or DPE can be compensated by the addition of an MTE. In addition, the MTE exhibits strong synergism with the TATA-box as well as the DPE. These findings indicate that the MTE is a novel downstream core promoter element that is important for transcription by RNA polymerase II. Publication Types: Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 15231738 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 8754800 # MOTIF: [JP] MA0038.1 Gfi ---------------------------------------------- Mol Cell Biol. 1996 Aug;16(8):4024-34. Gfi-1 encodes a nuclear zinc finger protein that binds DNA and functions as a transcriptional repressor. Zweidler-Mckay PA, Grimes HL, Flubacher MM, Tsichlis PN. Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA. The Gfi-1 proto-oncogene encodes a zinc finger protein with six C2H2-type, C-terminal zinc finger motifs and is activated by provirus integration in T-cell lymphoma lines selected for interleukin-2 independence in culture and in primary retrovirus-induced thymomas. Gfi-1 expression in adult animals is restricted to the thymus, spleen, and testis and is enhanced in mitogen-stimulated splenocytes. In this report, we show that Gfi-1 is a 55-kDa nuclear protein that binds DNA in a sequence-specific manner. The Gfi-1 binding site, TAAATCAC(A/T)GCA, was defined via random oligonucleotide selection utilizing a bacterially expressed glutathione S-transferase-Gfi-1 fusion protein. Binding to this site was confirmed by electrophoretic mobility shift assays and DNase I footprinting. Methylation interference analysis and electrophoretic mobility shift assays with mutant oliginucleotides defined the relative importance of specific bases at the consensus binding site. Deletion of individual zinc fingers demonstrated that only zinc fingers 3, 4, and 5 are required for sequence-specific DNA binding. Potential Gfi-1 binding sites were detected in a large number of eukaryotic promoter-enhancers, including the enhancers of several proto-oncogenes and cytokine genes and the enhancer of the human cytomegalovirus (HCMV) major immediate-early promoter, which contains two such sites. HCMV major immediate-early-chloramphenicol acetyltransferase reporter constructs, transfected into NIH 3T3 fibroblasts, were repressed by Gfi-1, and the repression was abrogated by mutation of critical residues in the two Gfi-1 binding sites. These results suggest that Gfi-1 may play a role in HCMV biology and may contribute to oncogenesis and T-cell activation by repressing the expression of genes that inhibit these processes. Publication Types: Comparative Study Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID: 8754800 [PubMed - indexed for MEDLINE] ---------------------------------------------- # MEDLINE ID: 9469818 # MOTIF: [JP] MA0060.1 NFYA ---------------------------------------------- Nucleic Acids Res. 1998 Mar 1;26(5):1135-43. A survey of 178 NF-Y binding CCAAT boxes. Mantovani R. Dipartimento di Genetica e Biologia dei Microrganismi, Universita di Milano, Via Celoria 26, 20133 Milano, Italy. mantor@imiucca.csi.unimi.it The CCAAT box is one of the most common elements in eukaryotic promoters, found in the forward or reverse orientation. Among the various DNA binding proteins that interact with this sequence, only NF-Y (CBF, HAP2/3/4/5) has been shown to absolutely require all 5 nt. Analysis of a database with 178 bona fide NF-Y binding sites in 96 unrelated promoters confirms this need and points to specific additional flanking nucleotides (C, Pu, Pu on the 5'-side and C/G, A/G, G,A/C, G on the 3'-side) required for efficient binding. The frequency of CCAAT boxes appears to be relatively higher in TATA-less promoters, particularly in the reverse ATTGG orientation. In TATA-containing promoters the CCAAT box is preferentially located in the -80/-100 region (mean position -89) and is not found nearer to the Start site than -50. In TATA-less promoters it is usually closer to the +1 signal (at -66 on average) and is sometimes present in proximity to the Cap site. The consensus and location of NF-Y binding sites parallel almost perfectly a previous general statistical study on CCAAT boxes in 502 unrelated promoters. This is an indication that NF-Y is the major, if not the sole, CCAAT box recognizing protein and that it might serve different roles in TATA-containing and TATA-less promoters. Publication Types: In Vitro Research Support, Non-U.S. Gov't Review PMID: 9469818 [PubMed - indexed for MEDLINE]