Functional divergence between the half-sites of the DNA-binding sequence for the yeast transcriptional regulator Rap1p

The yeast transcriptional regulator Rap1p binds to the DNA consensus sequence ACACCCAYACAYYY. We have previously shown that DNA-binding sites in which all four Y (Y = T or C) positions were Ts (UASrpg sequences) synergized more efficiently to activate transcription than sequences in which all Ys were Cs (telomere sequences) [F.-Z. Idrissi, J. Fernández-Larrea and B. Piña (1998) J. Mol. Biol. 284, 925-935]. Here we provide evidence that the DNA consensus sequence for Rap1p behaves as a combination of two ACAYYY half-sites with different functionality, the presence of Ts in the second half-site being the determinant for the transcriptional behaviour of the UASrpg sequences. DNA structure in the different complexes with Rap1p varied from being relatively uniform to appear rather distorted, this also being dependent on the presence of Ts in the second half-site. These distortions did not cause sharp bends or kinks in the DNA molecule. Computer analysis suggests that high-affinity binding of Rap1p to UASrpg sequences requires a rearrangement of the C-terminal Myb domain of the protein. We propose that the structural alterations in Rap1p-DNA complexes, both in the DNA and in the protein, affect the transcription potential of the complex in an allosteric manner. We also propose that the dimeric nature of the Rap1 DNA-binding domain is a key structural feature that explains the disparate functions of its DNA-binding sites in vivo.

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An effective approach for identification of in vivo protein-DNA binding sites from paired-end ChIP-Seq data

BMC Bioinformatics ◽

10.1186/1471-2105-11-81 ◽

2010 ◽

Vol 11 (1) ◽

pp. 81 ◽

Cited By ~ 16

Author(s):

Congmao Wang ◽

Jie Xu ◽

Dasheng Zhang ◽

Zoe A Wilson ◽

Dabing Zhang

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Dna Binding Sites

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Accurate and sensitive quantification of protein-DNA binding affinity

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1714376115 ◽

2018 ◽

Vol 115 (16) ◽

pp. E3692-E3701 ◽

Cited By ~ 32

Author(s):

Chaitanya Rastogi ◽

H. Tomas Rube ◽

Judith F. Kribelbauer ◽

Justin Crocker ◽

Ryan E. Loker ◽

...

Keyword(s):

Gene Expression ◽

Dna Binding ◽

Binding Sites ◽

Biophysical Model ◽

Regulatory Sequences ◽

Binding Modes ◽

Perfect Agreement ◽

Dna Binding Sites

Transcription factors (TFs) control gene expression by binding to genomic DNA in a sequence-specific manner. Mutations in TF binding sites are increasingly found to be associated with human disease, yet we currently lack robust methods to predict these sites. Here, we developed a versatile maximum likelihood framework named No Read Left Behind (NRLB) that infers a biophysical model of protein-DNA recognition across the full affinity range from a library of in vitro selected DNA binding sites. NRLB predicts human Max homodimer binding in near-perfect agreement with existing low-throughput measurements. It can capture the specificity of the p53 tetramer and distinguish multiple binding modes within a single sample. Additionally, we confirm that newly identified low-affinity enhancer binding sites are functional in vivo, and that their contribution to gene expression matches their predicted affinity. Our results establish a powerful paradigm for identifying protein binding sites and interpreting gene regulatory sequences in eukaryotic genomes.

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Identification and characterization of preferred DNA-binding sites for the Thermus thermophilus transcriptional regulator FadR

PLoS ONE ◽

10.1371/journal.pone.0184796 ◽

2017 ◽

Vol 12 (9) ◽

pp. e0184796 ◽

Cited By ~ 2

Author(s):

Minwoo Lee ◽

Hyejin Um ◽

Michael W. Van Dyke

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Thermus Thermophilus ◽

Transcriptional Regulator ◽

Dna Binding Sites ◽

Identification And Characterization

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Selection of DNA Binding Sites for Human Transcriptional Regulator GATA-6

Biochemical and Biophysical Research Communications ◽

10.1006/bbrc.1998.9374 ◽

1998 ◽

Vol 250 (3) ◽

pp. 682-688 ◽

Cited By ~ 38

Author(s):

Yuko Sakai ◽

Reiko Nakagawa ◽

Ryuichiro Sato ◽

Masatomo Maeda

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Transcriptional Regulator ◽

Dna Binding Sites ◽

Selection Of

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An A257V Mutation in the Bacillus subtilis Response Regulator Spo0A Prevents Regulated Expression of Promoters with Low-Consensus Binding Sites

Journal of Bacteriology ◽

10.1128/jb.00590-09 ◽

2009 ◽

Vol 191 (17) ◽

pp. 5489-5498 ◽

Cited By ~ 3

Author(s):

Steve D. Seredick ◽

Barbara M. Seredick ◽

David Baker ◽

George B. Spiegelman

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Transcription Initiation ◽

Molecular Basis ◽

Response Regulator ◽

Mutant Protein ◽

Wild Type ◽

Dna Binding Sites

ABSTRACT In Bacillus species, the master regulator of sporulation is Spo0A. Spo0A functions by both activating and repressing transcription initiation from target promoters that contain 0A boxes, the binding sites for Spo0A. Several classes of spo0A mutants have been isolated, and the molecular basis for their phenotypes has been determined. However, the molecular basis of the Spo0A(A257V) substitution, representative of an unusual phenotypic class, is not understood. Spo0A(A257V) is unusual in that it abolishes sporulation; in vivo, it fails to activate transcription from key stage II promoters yet retains the ability to repress the abrB promoter. To determine how Spo0A(A257V) retains the ability to repress but not stimulate transcription, we performed a series of in vitro and in vivo assays. We found unexpectedly that the mutant protein both stimulated transcription from the spoIIG promoter and repressed transcription from the abrB promoter, albeit twofold less than the wild type. A DNA binding analysis of Spo0A(A257V) showed that the mutant protein was less able to tolerate alterations in the sequence and arrangement of its DNA binding sites than the wild-type protein. In addition, we found that Spo0A(A257V) could stimulate transcription of a mutant spoIIG promoter in vivo in which low-consensus binding sites were replaced by high-consensus binding sites. We conclude that Spo0A(A257V) is able to bind to and regulate the expression of only genes whose promoters contain high-consensus binding sites and that this effect is sufficient to explain the observed sporulation defect.

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A micro-evolutionary change in target binding sites as key determinant of Ultrabithorax function in Drosophila.

10.1101/2021.08.17.456507 ◽

2021 ◽

Author(s):

Soumen Khan ◽

Saurabh J. Pradhan ◽

Guillaume Giraud ◽

Françoise Bleicher ◽

Rachel Paul ◽

...

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Morphological Changes ◽

Binding Motif ◽

Core Sequence ◽

Dna Binding Sites ◽

Target Binding ◽

Hox Protein

All Hox proteins are known to recognize, in vitro, similar DNA-binding sites containing a TAAT core sequence. This poor DNA-binding specificity is in sharp contrast with their specific functions in vivo. Here we report a new binding motif with TAAAT core sequence to which the Hox protein Ultrabithorax (Ubx) binds with higher affinity and specificity. Using transgenic and luciferase assays, we show that this new motif is critical for Ubx-mediated regulation of a target gene in Drosophila melanogaster. Interestingly, this new motif with TAAAT core sequences is not associated with the targets of Ubx in the honeybee, Apis mellifera, wherein hindwings are nearly identical to the forewings. We show that introduction of TAAAT motif in the place of TAAT motif is sufficient to bring an enhancer of a wing-promoting gene of A. mellifera under the regulation of Ubx. Our results, thus, suggest that binding motifs with a TAAAT core sequence may help identify functionally relevant direct targets of Ubx in D. melanogaster and the emergence of these binding sites may be crucial for Hox-mediated morphological changes during insect evolution.

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Identifying Sites Bound by Epstein-Barr Virus Nuclear Antigen 1 (EBNA1) in the Human Genome: Defining a Position-Weighted Matrix To Predict Sites Bound by EBNA1 in Viral Genomes

Journal of Virology ◽

10.1128/jvi.01974-08 ◽

2009 ◽

Vol 83 (7) ◽

pp. 2930-2940 ◽

Cited By ~ 45

Author(s):

Lindsay R. Dresang ◽

David T. Vereide ◽

Bill Sugden

Keyword(s):

Dna Binding ◽

Human Genome ◽

Binding Sites ◽

Epstein Barr Virus ◽

Consensus Sequence ◽

Nuclear Antigen ◽

Barr Virus ◽

Dna Binding Sites ◽

Weighted Matrix ◽

Epstein Barr

ABSTRACT We identified binding sites for Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) in the human genome using chromatin immunoprecipitation and microarrays. The sequences for these newly identified sites were used to generate a position-weighted matrix (PWM) for EBNA1's DNA-binding sites. This PWM helped identify additional DNA-binding sites for EBNA1 in the genomes of EBV, Kaposi's sarcoma-associated herpesvirus, and cercopithecine herpesvirus 15 (CeHV-15) (also called herpesvirus papio 15). In particular, a homologue of the Rep* locus in EBV was predicted in the genome of CeHV-15, which is notable because Rep* of EBV was not predicted by the previously developed consensus sequence for EBNA1's binding DNA. The Rep* of CeHV-15 functions as an origin of DNA synthesis in the EBV-positive cell line Raji; this finding thus builds on a set of DNA-binding sites for EBNA1 predicted in silico.

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Transcriptional repression by Msx-1 does not require homeodomain DNA-binding sites.

Molecular and Cellular Biology ◽

10.1128/mcb.15.2.861 ◽

1995 ◽

Vol 15 (2) ◽

pp. 861-871 ◽

Cited By ~ 110

Author(s):

K M Catron ◽

H Zhang ◽

S C Marshall ◽

J A Inostroza ◽

J M Wilson ◽

...

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Transcriptional Repression ◽

Binding Domain ◽

Specific Cell ◽

Homeodomain Protein ◽

Dna Binding Domain ◽

Dna Binding Sites

This study investigates the transcriptional properties of Msx-1, a murine homeodomain protein which has been proposed to play a key role in regulating the differentiation and/or proliferation state of specific cell populations during embryogenesis. We show, using basal and activated transcription templates, that Msx-1 is a potent repressor of transcription and can function through both TATA-containing and TATA-less promoters. Moreover, repression in vivo and in vitro occurs in the absence of DNA-binding sites for the Msx-1 homeodomain. Utilizing a series of truncated Msx-1 polypeptides, we show that multiple regions of Msx-1 contribute to repression, and these are rich in alanine, glycine, and proline residues. When fused to a heterologous DNA-binding domain, both N- and C-terminal regions of Msx-1 retain repressor function, which is dependent upon the presence of the heterologous DNA-binding site. Moreover, a polypeptide consisting of the full-length Msx-1 fused to a heterologous DNA-binding domain is a more potent repressor than either the N- or C-terminal regions alone, and this fusion retains the ability to repress transcription in the absence of the heterologous DNA site. We further show that Msx-1 represses transcription in vitro in a purified reconstituted assay system and interacts with protein complexes composed of TBP and TFIIA (DA) and TBP, TFIIA, and TFIIB (DAB) in gel retardation assays, suggesting that the mechanism of repression is mediated through interaction(s) with a component(s) of the core transcription complex. We speculate that the repressor function of Msx-1 is critical for its proposed role in embryogenesis as a regulator of cellular differentiation.

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Combinatorial bZIP dimers display complex DNA-binding specificity landscapes

eLife ◽

10.7554/elife.19272 ◽

2017 ◽

Vol 6 ◽

Cited By ~ 42

Author(s):

José A Rodríguez-Martínez ◽

Aaron W Reinke ◽

Devesh Bhimsaria ◽

Amy E Keating ◽

Aseem Z Ansari

Keyword(s):

Dna Binding ◽

Binding Sites ◽

Binding Specificity ◽

Human Cell Lines ◽

Protein Dimerization ◽

Site Preferences ◽

Dna Binding Specificity ◽

Genome Wide ◽

Half Site

How transcription factor dimerization impacts DNA-binding specificity is poorly understood. Guided by protein dimerization properties, we examined DNA binding specificities of 270 human bZIP pairs. DNA interactomes of 80 heterodimers and 22 homodimers revealed that 72% of heterodimer motifs correspond to conjoined half-sites preferred by partnering monomers. Remarkably, the remaining motifs are composed of variably-spaced half-sites (12%) or ‘emergent’ sites (16%) that cannot be readily inferred from half-site preferences of partnering monomers. These binding sites were biochemically validated by EMSA-FRET analysis and validated in vivo by ChIP-seq data from human cell lines. Focusing on ATF3, we observed distinct cognate site preferences conferred by different bZIP partners, and demonstrated that genome-wide binding of ATF3 is best explained by considering many dimers in which it participates. Importantly, our compendium of bZIP-DNA interactomes predicted bZIP binding to 156 disease associated SNPs, of which only 20 were previously annotated with known bZIP motifs.

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