CTCF mediates chromatin looping via N-terminal domain-dependent cohesin retention

Pugacheva, Elena M.; Kubo, Naoki; Loukinov, Dmitri; Tajmul, Md-D.; Kang, Sungyun; Kovalchuk, Alexander L.; Strunnikov, Alexander; Zentner, Gabriel E.; Ren, Bing; Lobanenkov, Victor

doi:10.1073/pnas.1911708117

Cited by 190 publications

(158 citation statements)

References 71 publications

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“…These co-immunoprecipitation data might not reflect a robust and stable interaction between CTCF and cohesin. Our observations are concordant with a parallel study demonstrating that several regions in the CTCF N terminus mediate cohesin retention, and that the CTCF N terminus is necessary for Hi-C peaks between TAD boundaries 20 .…”

Section: Rad21-halotag Kymographsupporting

confidence: 91%

“…Therefore, proper retention of cohesin at CTCF sites requires N(1-265), indicating that the CTCF N terminus either participates in inhibiting cohesin translocation (thereby promoting insulation) or-nonexclusively-protects blocked cohesin from unloading (thereby bolstering 5C peaks between CTCF sites). These observations are in line with a parallel study concluding that the N terminus is required for RAD21 occupancy at CTCF sites 20 . Given that deleting the CTCF N terminus led to milder insulation defects than complete CTCF depletion, and that deleting the C terminus had little-to-no effect, the ZF array mediates some degree of insulation and must therefore participate in halting cohesin translocation.…”

Section: Rad21-halotag Kymographsupporting

confidence: 91%

“…Through systematic truncations of the N terminus, we uncovered several regions important for genome folding and discovered a short protein motif that is both necessary and sufficient to recruit the PDS5A subunit of cohesin in a three-hybrid system. The PDS5Ainteracting region of CTCF is distinct from the N-terminal region recently reported to interact with RAD21-SA2 in vitro 19 and required for cohesin enrichment at CTCF sites 19,20 . This CTCF motif displays homology with the PDS5-binding domain of both WAPL and its competitors SORORIN and HASPIN.…”

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confidence: 87%

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Molecular basis of CTCF binding polarity in genome folding

et al. 2020

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Current models propose that boundaries of mammalian topologically associating domains (TADs) arise from the ability of the CTCF protein to stop extrusion of chromatin loops by cohesin. While the orientation of CTCF motifs determines which pairs of CTCF sites preferentially stabilize loops, the molecular basis of this polarity remains unclear. By combining ChIP-seq and single molecule live imaging we report that CTCF positions cohesin, but does not control its overall binding dynamics on chromatin. Using an inducible complementation system, we find that CTCF mutants lacking the N-terminus cannot insulate TADs properly. Cohesin remains at CTCF sites in this mutant, albeit with reduced enrichment. Given the orientation of CTCF motifs presents the N-terminus towards cohesin as it translocates from the interior of TADs, these observations explain how the orientation of CTCF binding sites translates into genome folding patterns.

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Section: Rad21-halotag Kymographsupporting

confidence: 91%

Section: Rad21-halotag Kymographsupporting

confidence: 91%

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confidence: 87%

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Molecular basis of CTCF binding polarity in genome folding

et al. 2020

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“…The involvement of ZNF143 in the establishment of CTCF loops (Wen et al 2018;Jung et al 2019) is supported by the increased strength of loops detected by SIP when CTCF and ZNF143 are both present at interacting anchors. The inability of CTCFL to form loops has been confirmed by a recent study that indicates that CTCFL cannot stop cohesin extrusion (Pugacheva et al 2020).…”

Section: Discussionmentioning

confidence: 78%

Analysis of Hi-C data using SIP effectively identifies loops in organisms from C. elegans to mammals

et al. 2020

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Chromatin loops are a major component of 3D nuclear organization, visually apparent as intense point-to-point interactions in Hi-C maps. Identification of these loops is a critical part of most Hi-C analyses. However, current methods often miss visually evident CTCF loops in Hi-C data sets from mammals, and they completely fail to identify high intensity loops in other organisms. We present SIP, Significant Interaction Peak caller, and SIPMeta, which are platform independent programs to identify and characterize these loops in a time- and memory-efficient manner. We show that SIP is resistant to noise and sequencing depth, and can be used to detect loops that were previously missed in human cells as well as loops in other organisms. SIPMeta corrects for a common visualization artifact by accounting for Manhattan distance to create average plots of Hi-C and HiChIP data. We then demonstrate that the use of SIP and SIPMeta can lead to biological insights by characterizing the contribution of several transcription factors to CTCF loop stability in human cells. We also annotate loops associated with the SMC component of the dosage compensation complex (DCC) in Caenorhabditis elegans and demonstrate that loop anchors represent bidirectional blocks for symmetrical loop extrusion. This is in contrast to the asymmetrical extrusion until unidirectional blockage by CTCF that is presumed to occur in mammals. Using HiChIP and multiway ligation events, we then show that DCC loops form a network of strong interactions that may contribute to X Chromosome–wide condensation in C. elegans hermaphrodites.

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“…An asterisk indicates an identical amino acid residue. ZFs(1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11) identified by MOTIF Search are indicated with colored boxes. SeeSupplementary Fig.…”

mentioning

confidence: 99%

Early vertebrate origin of CTCFL, a CTCF paralog, revealed by proximity-guided shark genome scaffolding

Kadota

Yamaguchi

Hara

et al. 2020

Sci Rep

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the nuclear protein ccctc-binding factor (ctcf) contributes as an insulator to chromatin organization in diverse animals. The gene encoding this protein has a paralog which was first identified to be expressed exclusively in the testis in mammals and designated as CTCFL (also called BORIS). CTCFL orthologs were reported only among amniotes, and thus CTCFL was once thought to have arisen in the amniote lineage. In this study, we identified elasmobranch CTCFL orthologs, and investigated its origin with the aid of a shark genome assembly improved by proximity-guided scaffolding. Our analysis employing evolutionary interpretation of syntenic gene location suggested an earlier timing of the gene duplication between CTCF and CTCFL than previously thought, that is, around the common ancestor of extant vertebrates. Also, our transcriptomic sequencing revealed a biased expression of the catshark CTCFL in the testis, suggesting the origin of the tissue-specific localization in mammals more than 400 million years ago. To understand the historical process of the functional consolidation of the long-standing chromatin regulator ctcf, its additional paralogs remaining in some of the descendant lineages for spatially restricted transcript distribution should be taken into consideration. The CCCTC-binding factor (CTCF) contains the C2H2 Zn finger-type DNA binding domains and plays a pivotal role in chromatin organization as an insulator in diverse metazoans 1,2. In vertebrates, the genome-wide binding landscape of the CTCF protein has been characterized for mammals 3,4 , sharks 5 , and the lamprey 6 , but the property of its paralog, CTCFL (also called BORIS, brother of the regulator of imprinted sites), has not been well characterized in a molecular phylogenetic context. CTCFL was first identified in human and mouse as a protein that functions in the testis and binds to the known target DNA of the CTCF protein in vitro 7. Their differential functions have been intensively investigated in germ cells and cancer cells mainly from epigenetic viewpoints 8-11. Comparison of amino acid sequences between CTCF and CTCFL exhibits a high similarity in the Zn finger DNA binding domain while the homology was low in other regions 7 , i.e., in the C-terminal region indispensable for the insulator function of CTCF 12. In contrast to the ubiquitously expressed CTCF 7 , the expression of CTCFL is restricted to the male testis, more specifically in the spermatocyte and the spermatogonia 7,13. Concordantly, while the mice lacking CTCF are embryonically lethal as early as E4.5 14 , mice lacking CTCFL are viable with phenotypes only in the testis, showing the marked reduction of its size caused in part by the increased rate of apoptosis during spermatogenesis 15. The report of CTCFL orthologs was long confined to mammals and lizards 16 but more recently the orthologs were identified in birds, turtle, snakes, and crocodiles 6,17. The gene expression patterns of CTCFL has been documented for only amniote species, and it is hypothesized that the testis...

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CTCF mediates chromatin looping via N-terminal domain-dependent cohesin retention

Cited by 190 publications

References 71 publications

Molecular basis of CTCF binding polarity in genome folding

Molecular basis of CTCF binding polarity in genome folding

Analysis of Hi-C data using SIP effectively identifies loops in organisms from C. elegans to mammals

Early vertebrate origin of CTCFL, a CTCF paralog, revealed by proximity-guided shark genome scaffolding

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