Polycomb repression of Hox genes involves spatial feedback but not domain compaction or demixing

Abstract

AbstractPolycomb group (PcG) proteins modulate higher-order genome folding and play a critical role in silencing transcription during development. It is commonly proposed that PcG dependent changes in genome folding, which compact chromatin, contribute directly to repression by blocking binding of activating complexes and demixing repressed targets from non-repressed chromatin. To test this model we utilized Optical Reconstruction of Chromatin Architecture (ORCA) to trace the 3-dimensional folding of theHoxagene cluster, a canonical Polycomb target, allowing us to analyze thousands of DNA traces in single cells. In cell types ranging from embryonic stem cells to brain tissue, we find that PcG-bound chromatin frequently explores decompact states and partial mixing with neighboring chromatin, while remaining uniformly repressed, challenging the repression-by-compaction model. Using polymer physics simulations, we show that the flexible ensembles we observe can be explained by dynamic contacts mediated by multivalent interactions that are too weak to induce phase separation. Instead, these transient contacts contribute to accurate propagation of the epigenetic state without ectopic spreading or gradual erosion. We propose that the distinctive 3D organization of Polycomb chromatin, reflects a mechanism of “spatial feedback” required for stable repression.