Context-dependent perturbations in chromatin folding and the transcriptome by cohesin and related factors [RNA-seq]

Cohesin plays vital roles in chromatin folding and gene expression regulation, cooperating with such factors as cohesin loaders, unloaders, and the insulation factor CTCF. Although models of regulation have been proposed (e.g., loop extrusion), how cohesin and related factors collectively or individually regulate the hierarchical chromatin structure and gene expression remains unclear. We have depleted cohesin and related factors and then conducted a comprehensive evaluation of the resulting 3D genome, transcriptome and epigenome data. We observed substantial variation in depletion effects among factors at topologically associating domain (TAD) boundaries and on interTAD interactions, which were related to epigenomic status. Total RNA was isolated using Trizol (Thermo Fisher Scientific) and a Nucleospin RNA kit (Macherey-Nagel). rRNA was removed with the Ribo-Zero Gold rRNA Removal Kit (Illumina), followed by sequencing library preparation with the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England Biolabs).