PAR-Driven Condensation Maintains Stalled Replication Fork Stability

Abstract

Poly(ADP-ribose) (PAR) is a nucleic acid-like heterogeneous polymer in nature. Recently, it was found to engage in liquid-liquid phase separation (LLPS), generating condensates as an emerging class of subcellular structures with pivotal functions in response to stimuli. As a post-translational modification catalyzed by PAR polymerases (PARPs), PAR is known to modulate many key events in cells. However, its involvement in biomolecular condensation remains elusive. Through an imaging-based screening of small molecules with diverse biological activities, we here discovered that PAR undergoes LLPS upon inhibiting proteasome in different types of cells, resulting in co-condensation of PAR with proteasome and ubiquitin chains in nucleus. This unprecedented co-condensation is dependent on PARP2 not PARP1 and requires K6-linked ubiquitylation. PAR is shown for the first time to directly interact with ubiquitin chains. Notably, stalled DNA replication forks arose from proteasome inhibition are co-localized with PAR-proteasome-ubiquitin chain condensates. By attenuating replication and stabilizing stalled replication forks, PAR-proteasome-ubiquitin chain condensates sustain genomic integrity under proteasomal stress. This work demonstrates a self-protective mechanism in stressed cells and provides fundamental understanding of PAR condensation in cell biology.