Re-evaluating the roles of cohesin and CTCF in genome folding

Animal genomes are organized into topologically associating domains (TADs). TADs are linked to important biological functions, including enhancer-mediated transcriptional regulation. How TADs form and what role they play in gene regulation is still widely researched and debated. Here, we used a microscopy approach, Optical Reconstruction of Chromatin Architecture (ORCA), to measure how cohesin and CTCF organize the genome and the consequences for gene regulation.

Previous work has shown that cohesin and CTCF are essential for TAD formation. However, despite a similar phenotype of TAD loss upon depletion of either protein, polymer models and experiments propose that cohesin and CTCF play different roles in genome organization. Cohesin is believed to facilitate intra-TAD interactions at the expense of inter-TAD contacts, while CTCF is thought to prevent inter-TAD mixing. Consequently, this would suggest a model where cohesin contributes to enhancer-promoter specificity between TADs while CTCF works to prevent erroneous contacts across TAD boundaries.

Our ORCA data challenges these models. Notably, we found that cohesin is required for maintaining proximity at the sub-3Mb scale and for decreasing ectopic contacts with more distal regions (>5Mb). CTCF organizes these loops along an axial/radial axis which favors long-range interactions among CTCF proximal sites and opposes them along distal sites. Our data provide a structural understanding of how cohesin and CTCF organize the genome at different genomic scales. Based on these measurements we propose a revised model of how cohesin and CTCF regulate enhancer-promoter specificity.