p97 Promotes a Conserved Mechanism of Helicase Unloading during DNA Cross-Link Repair

Abstract

Inter-strand crosslinks (ICLs) are extremely toxic DNA lesions that create an impassable roadblock to DNA replication. When a replication fork collides with an ICL, it triggers a damage response that promotes multiple DNA processing events required to excise the crosslink from chromatin and resolve the stalled replication fork. One of the first steps in this process involves displacement of the CMG replicative helicase (comprised of Cdc45, MCM2-7, and GINS), which obstructs the underlying crosslink. Here we report that the p97/Cdc48/VCP segregase plays a critical role in ICL repair by unloading the CMG complex from chromatin. Eviction of the stalled helicase involves K48-linked polyubiquitylation of MCM7, p97-mediated extraction of CMG, and a largely degradation-independent mechanism of MCM7 de-ubiquitylation. Our results show that ICL repair and replication termination both utilize a similar mechanism to displace the CMG complex from chromatin. However, unlike termination, repair-mediated helicase unloading involves the tumor suppressor protein BRCA1, which acts upstream of MCM7 ubiquitylation and p97 recruitment. Together, these findings indicate that p97 plays a conserved role in dismantling the CMG helicase complex during different cellular events, but that distinct regulatory signals ultimately control when and where unloading takes place.