CTCF: the protein, the binding partners, the binding sites and their chromatin loops

CTCF has it all. The transcription factor binds to tens of thousands of genomic sites, some tissue-specific, others ultra-conserved. It can act as a transcriptional activator, repressor and insulator, and it can pause transcription. CTCF binds at chromatin domain boundaries, at enhancers and gene promoters, and inside gene bodies. It can attract many other transcription factors to chromatin, including tissue-specific transcriptional activators, repressors, cohesin and RNA polymerase II, and it forms chromatin loops. Yet, or perhaps therefore, CTCF's exact function at a given genomic site is unpredictable. It appears to be determined by the associated transcription factors, by the location of the binding site relative to the transcriptional start site of a gene, and by the site's engagement in chromatin loops with other CTCF-binding sites, enhancers or gene promoters. Here, we will discuss genome-wide features of CTCF binding events, as well as locus-specific functions of this remarkable transcription factor.

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HOT or not: examining the basis of high-occupancy target regions

10.1101/107680 ◽

2017 ◽

Cited By ~ 3

Author(s):

Katarzyna Wreczycka ◽

Vedran Franke ◽

Bora Uyar ◽

Ricardo Wurmus ◽

Altuna Akalin

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Binding Sites ◽

Cell Types ◽

Data Sets ◽

Gene Promoters ◽

Quadruplex Dna ◽

Golden Standard ◽

Multiple Cell ◽

Multiple Species

AbstractHigh-occupancy target (HOT) regions are the segments of the genome with unusually high number of transcription factor binding sites. These regions are observed in multiple species and thought to have biological importance due to high transcription factor occupancy. Furthermore, they coincide with house-keeping gene promoters and the associated genes are stably expressed across multiple cell types. Despite these features, HOT regions are solemnly defined using ChIP-seq experiments and shown to lack canonical motifs for transcription factors that are thought to be bound there. Although, ChIP-seq experiments are the golden standard for finding genome-wide binding sites of a protein, they are not noise free. Here, we show that HOT regions are likely to be ChIP-seq artifacts and they are similar to previously proposed “hyper-ChIPable” regions. Using ChIP-seq data sets for knocked-out transcription factors, we demonstrate presence of false positive signals on HOT regions. We observe sequence characteristics and genomic features that are discriminatory of HOT regions, such as GC/CpG-rich k-mers and enrichment of RNA-DNA hybrids (R-loops) and DNA tertiary structures (G-quadruplex DNA). The artificial ChIP-seq enrichment on HOT regions could be associated to these discriminatory features. Furthermore, we propose strategies to deal with such artifacts for the future ChIP-seq studies.

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Genome-wide targeting of the epigenetic regulatory protein CTCF to gene promoters by the transcription factor TFII-I

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1416674112 ◽

2015 ◽

Vol 112 (7) ◽

pp. E677-E686 ◽

Cited By ~ 39

Author(s):

Rodrigo Peña-Hernández ◽

Maud Marques ◽

Khalid Hilmi ◽

Teijun Zhao ◽

Amine Saad ◽

...

Keyword(s):

Transcription Factor ◽

Rna Polymerase Ii ◽

Transcription Initiation ◽

Cellular Response ◽

Target Genes ◽

Regulatory Protein ◽

Metabolic Stress ◽

Ctcf Binding ◽

Gene Promoters ◽

The Impact

CCCTC-binding factor (CTCF) is a key regulator of nuclear chromatin structure and gene regulation. The impact of CTCF on transcriptional output is highly varied, ranging from repression to transcriptional pausing and transactivation. The multifunctional nature of CTCF may be directed solely through remodeling chromatin architecture. However, another hypothesis is that the multifunctional nature of CTCF is mediated, in part, through differential association with protein partners having unique functions. Consistent with this hypothesis, our mass spectrometry analyses of CTCF interacting partners reveal a previously undefined association with the transcription factor general transcription factor II-I (TFII-I). Biochemical fractionation of CTCF indicates that a distinct CTCF complex incorporating TFII-I is assembled on DNA. Unexpectedly, we found that the interaction between CTCF and TFII-I is essential for directing CTCF to the promoter proximal regulatory regions of target genes across the genome, particularly at genes involved in metabolism. At genes coregulated by CTCF and TFII-I, we find knockdown of TFII-I results in diminished CTCF binding, lack of cyclin-dependent kinase 8 (CDK8) recruitment, and an attenuation of RNA polymerase II phosphorylation at serine 5. Phenotypically, knockdown of TFII-I alters the cellular response to metabolic stress. Our data indicate that TFII-I directs CTCF binding to target genes, and in turn the two proteins cooperate to recruit CDK8 and enhance transcription initiation.

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Rfx2 stabilizes Foxj1 binding at chromatin loops to enable multiciliated cell gene expression

10.1101/085571 ◽

2016 ◽

Cited By ~ 2

Author(s):

Ian K Quigley ◽

Chris Kintner

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Transcription Factors ◽

Target Genes ◽

Gene Promoters ◽

Topological Domains ◽

Chromatin Loops ◽

Organ Systems ◽

Cilia Formation ◽

Motile Cilia

AbstractCooperative transcription factor binding at cis-regulatory sites in the genome drives robust eukaryotic gene expression, and many such sites must be coordinated to produce coherent transcriptional programs. The transcriptional program leading to motile cilia formation requires members of the DNA-binding forkhead (Fox) and Rfx transcription factor families and these factors co-localize to cilia gene promoters, but it is not clear how many cilia genes are regulated by these two factors, whether these factors act directly or indirectly, or how these factors act with specificity in the context of a 3-dimensional genome. Here, we use genome-wide approaches to show that cilia genes reside at the boundaries of topological domains and that these areas have low enhancer density. We show that the transcription factors Foxj1 and Rfx2 binding occurs in the promoters of more cilia genes than other known cilia transcription factors and that while Rfx2 binds directly to promoters and enhancers equally, Foxj1 prefers direct binding to enhancers and is stabilized at promoters by Rfx2. Finally, we show that Rfx2 and Foxj1 lie at the anchor endpoints of chromatin loops, suggesting that target genes are activated when Foxj1 bound at distal sites is recruited via a loop created by Rfx2 binding at both sites. We speculate that the primary function of Rfx2 is to stabilize distal enhancers with proximal promoters by operating as a scaffolding factor, bringing key regulatory domains bound by Foxj1 into close physical proximity and enabling coordinated cilia gene expression.Author SummaryThe multiciliated cell extends hundreds of motile cilia to produce fluid flow in the airways and other organ systems. The formation of this specialized cell type requires the coordinated expression of hundreds of genes in order to produce all the protein parts motile cilia require. While a relatively small number of transcription factors has been identified that promote gene expression during multiciliate cell differentiation, it is not clear how they work together to coordinate the expression of genes required for multiple motile ciliation. Here, we show that two transcription factors known to drive cilia formation, Foxj1 and Rfx2, play complementary roles wherein Foxj1 activates target genes but tends not to bind near them in the genome, whereas Rfx2 can’t activate target genes by itself but instead acts as a scaffold by localizing Foxj1 to the proper targets. These results suggest not only a mechanism by which complex gene expression is coordinated in multiciliated cells, but also how transcriptional programs in general could be modular and deployed across different cellular contexts with the same basic promoter configuration.

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Persistent Interactions of Core Histone Tails with Nucleosomal DNA following Acetylation and Transcription Factor Binding

Molecular and Cellular Biology ◽

10.1128/mcb.18.11.6293 ◽

1998 ◽

Vol 18 (11) ◽

pp. 6293-6304 ◽

Cited By ~ 96

Author(s):

Vesco Mutskov ◽

Delphine Gerber ◽

Dimitri Angelov ◽

Juan Ausio ◽

Jerry Workman ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Molecular Biology ◽

Binding Sites ◽

Cross Linking ◽

Uv Laser ◽

Histone Tail ◽

Histone Tails ◽

Nucleosomal Dna ◽

Core Histones

ABSTRACT In this study, we examined the effect of acetylation of the NH2 tails of core histones on their binding to nucleosomal DNA in the absence or presence of bound transcription factors. To do this, we used a novel UV laser-induced protein-DNA cross-linking technique, combined with immunochemical and molecular biology approaches. Nucleosomes containing one or five GAL4 binding sites were reconstituted with hypoacetylated or hyperacetylated core histones. Within these reconstituted particles, UV laser-induced histone-DNA cross-linking was found to occur only via the nonstructured histone tails and thus presented a unique tool for studying histone tail interactions with nucleosomal DNA. Importantly, these studies demonstrated that the NH2 tails were not released from nucleosomal DNA upon histone acetylation, although some weakening of their interactions was observed at elevated ionic strengths. Moreover, the binding of up to five GAL4-AH dimers to nucleosomes occupying the central 90 bp occurred without displacement of the histone NH2 tails from DNA. GAL4-AH binding perturbed the interaction of each histone tail with nucleosomal DNA to different degrees. However, in all cases, greater than 50% of the interactions between the histone tails and DNA was retained upon GAL4-AH binding, even if the tails were highly acetylated. These data illustrate an interaction of acetylated or nonacetylated histone tails with DNA that persists in the presence of simultaneously bound transcription factors.

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Specific transcription factors stimulate simian virus 40 and polyomavirus origins of DNA replication

Molecular and Cellular Biology ◽

10.1128/mcb.12.6.2514-2524.1992 ◽

1992 ◽

Vol 12 (6) ◽

pp. 2514-2524 ◽

Cited By ~ 1

Author(s):

Z S Guo ◽

M L DePamphilis

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Dna Replication ◽

Binding Sites ◽

Simian Virus 40 ◽

Simian Virus ◽

T Antigen ◽

Activation Domains ◽

Auxiliary Component ◽

Cell Extracts

The origins of DNA replication (ori) in simian virus 40 (SV40) and polyomavirus (Py) contain an auxiliary component (aux-2) composed of multiple transcription factor binding sites. To determine whether this component stimulated replication by binding specific transcription factors, aux-2 was replaced by synthetic oligonucleotides that bound a single transcription factor. Sp1 and T-antigen (T-ag) sites, which exist in the natural SV40 aux-2 sequence, provided approximately 75 and approximately 20%, respectively, of aux-2 activity when transfected into monkey cells. In cell extracts, only T-ag sites were active. AP1 binding sites could replace completely either SV40 or Py aux-2. Mutations that eliminated AP1 binding also eliminated AP1 stimulation of replication. Yeast GAL4 binding sites that strongly stimulated transcription in the presence of GAL4 proteins failed to stimulate SV40 DNA replication, although they did partially replace Py aux-2. Stimulation required the presence of proteins consisting of the GAL4 DNA binding domain fused to specific activation domains such as VP16 or c-Jun. These data demonstrate a clear role for transcription factors with specific activation domains in activating both SV40 and Py ori. However, no correlation was observed between the ability of specific proteins to stimulate promoter activity and their ability to stimulate origin activity. We propose that only transcription factors whose specific activation domains can interact with the T-ag initiation complex can stimulate SV40 and Py ori-core activity.

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TFutils: Data structures for transcription factor bioinformatics

F1000Research ◽

10.12688/f1000research.17976.1 ◽

2019 ◽

Vol 8 ◽

pp. 152

Author(s):

Benjamin J. Stubbs ◽

Shweta Gopaulakrishnan ◽

Kimberly Glass ◽

Nathalie Pochet ◽

Celine Everaert ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Data Structures ◽

Binding Sites ◽

Genome Wide Association Study ◽

Genome Wide Association ◽

Bioconductor Package ◽

Genome Wide ◽

Study Results ◽

Integrative Analyses

DNA transcription is intrinsically complex. Bioinformatic work with transcription factors (TFs) is complicated by a multiplicity of data resources and annotations. The Bioconductor package TFutils includes data structures and functions to enhance the precision and utility of integrative analyses that have components involving TFs. TFutils provides catalogs of human TFs from three reference sources (CISBP, HOCOMOCO, and GO), a catalog of TF targets derived from MSigDb, and multiple approaches to enumerating TF binding sites. Aspects of integration of TF binding patterns and genome-wide association study results are explored in examples.

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Virtual ChIP-seq: predicting transcription factor binding by learning from the transcriptome

10.1101/168419 ◽

2018 ◽

Cited By ~ 10

Author(s):

Mehran Karimzadeh ◽

Michael M. Hoffman

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Binding Sites ◽

Cell Types ◽

Transcription Factor Binding ◽

Regulatory Function ◽

Factor Binding ◽

Link Type ◽

Genomic Regions ◽

Factor Sequence

AbstractMotivationIdentifying transcription factor binding sites is the first step in pinpointing non-coding mutations that disrupt the regulatory function of transcription factors and promote disease. ChIP-seq is the most common method for identifying binding sites, but performing it on patient samples is hampered by the amount of available biological material and the cost of the experiment. Existing methods for computational prediction of regulatory elements primarily predict binding in genomic regions with sequence similarity to known transcription factor sequence preferences. This has limited efficacy since most binding sites do not resemble known transcription factor sequence motifs, and many transcription factors are not even sequence-specific.ResultsWe developed Virtual ChIP-seq, which predicts binding of individual transcription factors in new cell types using an artificial neural network that integrates ChIP-seq results from other cell types and chromatin accessibility data in the new cell type. Virtual ChIP-seq also uses learned associations between gene expression and transcription factor binding at specific genomic regions. This approach outperforms methods that predict TF binding solely based on sequence preference, pre-dicting binding for 36 transcription factors (Matthews correlation coefficient > 0.3).AvailabilityThe datasets we used for training and validation are available at https://virchip.hoffmanlab.org. We have deposited in Zenodo the current version of our software (http://doi.org/10.5281/zenodo.1066928), datasets (http://doi.org/10.5281/zenodo.823297), predictions for 36 transcription factors on Roadmap Epigenomics cell types (http://doi.org/10.5281/zenodo.1455759), and predictions in Cistrome as well as ENCODE-DREAM in vivo TF Binding Site Prediction Challenge (http://doi.org/10.5281/zenodo.1209308).

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Holocentromeres are dispersed point centromeres localized at transcription factor hotspots

eLife ◽

10.7554/elife.02025 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 61

Author(s):

Florian A Steiner ◽

Steven Henikoff

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Caenorhabditis Elegans ◽

Binding Sites ◽

Specific Binding ◽

Basic Unit ◽

Centromeric Chromatin ◽

Sequence Composition ◽

Specific Binding Sites ◽

Mechanistic Basis

Centromeres vary greatly in size and sequence composition, ranging from ‘point’ centromeres with a single cenH3-containing nucleosome to ‘regional’ centromeres embedded in tandemly repeated sequences to holocentromeres that extend along the length of entire chromosomes. Point centromeres are defined by sequence, whereas regional and holocentromeres are epigenetically defined by the location of cenH3-containing nucleosomes. In this study, we show that Caenorhabditis elegans holocentromeres are organized as dispersed but discretely localized point centromeres, each forming a single cenH3-containing nucleosome. These centromeric sites co-localize with kinetochore components, and their occupancy is dependent on the cenH3 loading machinery. These sites coincide with non-specific binding sites for multiple transcription factors (‘HOT’ sites), which become occupied when cenH3 is lost. Our results show that the point centromere is the basic unit of holocentric organization in support of the classical polycentric model for holocentromeres, and provide a mechanistic basis for understanding how centromeric chromatin might be maintained.

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The Pu.1 Locus Is Differentially Regulated at the Level of Chromatin Structure and Noncoding Transcription by Alternate Mechanisms at Distinct Developmental Stages of Hematopoiesis

Molecular and Cellular Biology ◽

10.1128/mcb.00905-07 ◽

2007 ◽

Vol 27 (21) ◽

pp. 7425-7438 ◽

Cited By ~ 40

Author(s):

Maarten Hoogenkamp ◽

Hanna Krysinska ◽

Richard Ingram ◽

Gang Huang ◽

Rachael Barlow ◽

...

Keyword(s):

T Cells ◽

Transcription Factor ◽

Transcription Factors ◽

B Cells ◽

Regulatory Element ◽

Precursor Cells ◽

Hematopoietic Stem ◽

Tissue Specific ◽

Specific Regulation

ABSTRACT The Ets family transcription factor PU.1 is crucial for the regulation of hematopoietic development. Pu.1 is activated in hematopoietic stem cells and is expressed in mast cells, B cells, granulocytes, and macrophages but is switched off in T cells. Many of the transcription factors regulating Pu.1 have been identified, but little is known about how they organize Pu.1 chromatin in development. We analyzed the Pu.1 promoter and the upstream regulatory element (URE) using in vivo footprinting and chromatin immunoprecipitation assays. In B cells, Pu.1 was bound by a set of transcription factors different from that in myeloid cells and adopted alternative chromatin architectures. In T cells, Pu.1 chromatin at the URE was open and the same transcription factor binding sites were occupied as in B cells. The transcription factor RUNX1 was bound to the URE in precursor cells, but binding was down-regulated in maturing cells. In PU.1 knockout precursor cells, the Ets factor Fli-1 compensated for the lack of PU.1, and both proteins could occupy a subset of Pu.1 cis elements in PU.1-expressing cells. In addition, we identified novel URE-derived noncoding transcripts subject to tissue-specific regulation. Our results provide important insights into how overlapping, but different, sets of transcription factors program tissue-specific chromatin structures in the hematopoietic system.

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