CUL2^LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle

Abstract

The eukaryotic replisome is disassembled in each cell cycle, dependent upon ubiquitylation of the CMG helicase. Studies of Saccharomyces cerevisiae, Caenorhabditis elegans and Xenopus laevis have revealed surprising evolutionary diversity in the ubiquitin ligases that control CMG ubiquitylation, but regulated disassembly of the mammalian replisome has yet to be explored. Here, we describe a model system for studying the ubiquitylation and chromatin extraction of the mammalian CMG replisome, based on mouse embryonic stem cells. We show that the ubiquitin ligase CUL2LRR1 is required for ubiquitylation of the CMG‐MCM7 subunit during S‐phase, leading to disassembly by the p97 ATPase. Moreover, a second pathway of CMG disassembly is activated during mitosis, dependent upon the TRAIP ubiquitin ligase that is mutated in primordial dwarfism and mis‐regulated in various cancers. These findings indicate that replisome disassembly in diverse metazoa is regulated by a conserved pair of ubiquitin ligases, distinct from those present in other eukaryotes.

MCM7 Ubiquitination Regulates CMG Helicase Disassembly In Human

“…Together, these results suggest that deubiquitylation of MCM7 mediated by p97 and proteasomal degradation could drive the disassembly of the replicative helicase during termination. Consistent with the results for S. cerevisiae, X. laevis, C. elegans embryos and mouse embryonic stem cells [7,8,13] , we identi ed that polyubiquitylation of MCM7 plays a conserved role in regulating replisome disassembly from chromatin in humans.…”

“…3A and 4F) have not been reported, which remains to be further investigated. In mouse embryonic stem cells, study found that ubiquitin ligases CUL2LRR1 and TRAIP control p97dependent replisome disassembly during DNA replication termination and mitosis, respectively [13] , which may also regulate the ubiquitination of MCM7 in Human, and is worth for further investigation.…”

“…Proteasome and Cdc48/p97 segregase are required for MCM7 deubiquitylation and CMG disassembly In X. laevis and S. cerevisiae, MCM7 deubiquitylation during replication termination depends on p97/Cdc48/VCP segregase but not proteasomal degradation [7,8] . MCM7 polyubiquitylation acts as the signal for p97-mediated extraction and unloading of the CMG complex from chromatin [7,8,10,13] . To investigate the mechanism of MCM7 deubiquitylation in human tumor cells, we inhibited proteasomal degradation with MG-132 treatment, which showed increased ubiquitination of MCM7 and prolonged CMG association on chromatin in the late S phase (Figure 6B).…”

“…When two replisomes converge at an interstrand DNA crosslink (ICL), TRAIP ubiquitylates MCM7 of CMG to activate CMG unloading and entry into the Fanconi anemia (FA) pathway in Xenopus egg extracts [12] . A recent study in mouse embryonic stem cells found that ubiquitin ligases CUL2LRR1 and TRAIP control p97-dependent replisome disassembly during DNA replication termination and mitosis, respectively [13] . However, the mechanism of human chromosomal replication dissociation is incompletely understood.…”

Traip Mitotic Function Controls Brain Size

“…A cell entering mitosis may have under-replicated sister chromatids (URSCs) which contain loci that failed to complete replication during interphase, and these URSCs must be resolved to prevent DNA bridges during anaphase. TRAIP initiates the resolution of URSCs by ubiquitylating the MCM7 subunit of CMG helicase, thus triggering replication machinery unloading from stalled forks and allowing sister chromatid resolution via mitotic DNA repair synthesis (Deng et al, 2019; Priego Moreno et al, 2019; Sonneville et al, 2019; Villa et al, 2021).…”