RB loss modulates chromatin organization by regulating cohesin-dependent loops and enhancer-promoter interactions

The retinoblastoma protein (RB) is a well-characterized repressor of E2F transcriptional activity that controls genes involved in cell proliferation during the G1 phase. Here, we examine the effect of RB on chromatin organization and uncover a non-canonical role for RB in which it promotes the removal of cohesin from insulators and thereby induces the expression of adjacent genes. We identify that RB colocalizes with cohesin across the human genome. During mitosis, RB facilitates the removal of cohesin from CTCF sites, with this effect persisting into early G1, impacting loop formation and extrusion at topologically associating domain (TAD) boundaries. Chromosome conformation capture assays reveal that RB reduces insulation at TAD boundaries and promotes enhancer-promoter interactions marked by histone 3 lysine 27 acetylation. In this way, RB enhances the transcription of hundreds of genes beyond the E2F transcriptional program, a finding that expands our understanding of this key tumor suppressor and cell cycle regulator.