Manipulation of Long-Range Chromatin Interactions through CTCF Phase Separation for Cell Fate Control

Gene expression is controlled under spatial chromatin structures with short-range in topologically associating domains (TAD) and long-range chromatin interactions between TADs, compartments or chromosomes, and disruption of chromatin structure leads to human diseases. The mechanism of short-range chromatin interactions has been well characterized by loop-extrusion model, but little is known about how long-range chromatin interactions are organized. Here, we demonstrate that CTCF contributes to long-range chromatin interactions via phase separation. Surprisingly, RYBP is required for the phase separation and long-range chromatin organization of CTCF. Artificial CTCF phase seperation restores the long-range chromatin interactions and corresponding gene expression which were eliminated by RYBP depletion, and manipulation of CTCF phase separation also maintains pluripotency and inhibits differentation of embryonic stem cells. These findings support a model that long-range chromatin interactions are organized through phase sepearation of architectural protein, and further reveals the distinct mechanisms of architectural protein in organizing short-range and long-range chromatin interactions. Examination of RNA-seq, Hi-C, CTCF HiChIP, RYBP HiCHIP NGS data before or/and after RYBP knock out.