Motif oriented high-resolution analysis of ChIP-seq data reveals the topological order of CTCF and cohesin proteins on DNA

Abstract

BackgroundChIP-seq provides a wealth of information on the approximate location of DNA-binding proteins genome-wide. It is known that the targeted motifs in most cases can be found at the peak centers. A high resolution mapping of ChIP-seq peaks could in principle allow the fine mapping of the protein constituents within protein complexes, but the current ChIP-seq analysis pipelines do not target the basepair resolution strand specific mapping of peak summits.ResultsThe approach proposed here is based on i) locating regions that are bound by a sufficient number of proteins constituting a complex; ii) determining the position of the underlying motif using either a direct or a de novo motif search approach; and iii) determining the exact location of the peak summits with respect to the binding motif in a strand specific manner. We applied this method for analyzing the CTCF/cohesin complex, which holds together DNA loops. The relative positions of the constituents of the complex were determined with one-basepair estimated accuracy. Mapping the positions on a 3D model of DNA made it possible to deduce the approximate local topology of the complex that allowed us to predict how the CTCF/cohesin complex locks the DNA loops. As the positioning of the proteins was not compatible with previous models of loop closure, we proposed a plausible “double embrace” model in which the DNA loop is held together by two adjacent cohesin rings in such a way that the ring anchored by CTCF to one DNA duplex encircles the other DNA double helix and vice versa.ConclusionsA motif-centered, strand specific analysis of ChIP-seq data improves the accuracy of determining peak positions. If a genome contains a large number of binding sites for a given protein complex, such as transcription factor heterodimers or transcription factor/cofactor complexes, the relative position of the constituent proteins on the DNA can be established with an accuracy that allow one to deduce the local topology of the protein complex. The proposed high resolution mapping approach of ChIP-seq data is applicable for detecting the contact topology of DNA-binding protein complexes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2940-7) contains supplementary material, which is available to authorized users.

Integrative characterization of G-Quadruplexes in the three-dimensional chromatin structure

“…We calculated the distance between G-quadruplexes/CTCF/RAD21/ SMC3 ChIP-seq peaks and forward/reverse CTCF motifs ( Figure S3A-B). In line with previous study, we found a characteristic shift pattern between CTCF motifs and cohesin protein ChIP-seq peaks [46]. Surprisingly, similar to cohesin protein ChIP-seq peaks, G-quadruplexes tend to distribute downstream of forward CTCF motifs (average distance = 3.15 bp, 2.18 bp, and 5.21 bp for G-quadruplexes, SMC3, and RAD21, respectively) and upstream of reverse CTCF motifs (average distance = −4.49 bp, −2.31 bp, and −5.79 bp for G-quadruplexes, SMC3, and RAD21, respectively).…”

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

“…In contrast, the C terminus of CTCF was dispensable for cohesin occupancy and loop formation, challenging previous work reporting that the C terminus of CTCF directly interacts with SA2, an external subunit of cohesin ring, and thus provides a bridge between CTCF and the cohesin complex (34). A role for the N terminus of CTCF in cohesin retention is consistent with high-resolution ChIP-seq analysis showing that cohesin binding tends to be shifted toward the N terminus of CTCF (37). Besides, an alternative form of CTCF missing its N terminus was shown to be unable to retain cohesin at CTCF binding sites (55).…”

“…However, precise deletion of the same peptide sequence from the C terminus of CTCF does not disrupt the interaction of CTCF with cohesin, as shown in two independent studies (35,36). Furthermore, a highresolution analysis of ChIP-seq data based on the orientation of -V5-TAG CTCF consensus sequences showed that cohesin occupancy at CTCF binding sites is usually shifted toward the N terminus rather than the C terminus of CTCF (26,37). To map the CTCF domain responsible for cohesin binding in vivo, we employed the mut CH12 cell system described above (Fig.…”

Functional roles of CTCF in breast cancer

“…CTCF-depletion studies identified the function of CTCF in association with chromatin structure in mouse embryonic stem cells ( 2 ). For maintaining stable genomic complex, CTCF and the cohesin complex, consisting of SMC3, SMC1, RAD21, and STAG1 or STAG2, can co-localize ( 13 ). Moreover, global analysis of CTCF, SMC3, and RAD1 shift-banding patterns have demonstrated the proximity of protein-DNA binding motif sequences ( 13 ).…”