Cell-Cycle-Regulated Interaction between Mcm10 and Double Hexameric Mcm2-7 Is Required for Helicase Splitting and Activation during S Phase

Quan, Yun; Xia, Yisui; Liu, Lu; Cui, Jiamin; Li, Zhen; Cao, Qinhong; Chen, Xiaojiang S.; Campbell, Judith L.; Lou, Huiqiang

doi:10.1016/j.celrep.2015.11.018

Cited by 57 publications

(71 citation statements)

References 45 publications

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“…1A). Consistent with recent findings (Quan et al 2015;Douglas and Diffley, 2016), mass spectrometry confirmed that these proteins were Mcm4 and Mcm6 and identified Mcm2 as the third protein (Supplemental Table S1). …”

Section: Mcm10 Binds a Conserved Region In The Mcm2 N-terminal Domainsupporting

confidence: 75%

“…Recruitment of Cdc45 or GINS to Mcm2-7 is independent of Mcm10 (Kanke et al 2012;van Deursen et al 2012;Watase et al 2012;Yeeles et al 2015). In contrast, Mcm10 is required for initial DNA unwinding at origins of replication (Kanke et al 2012;van Deursen et al 2012;Watase et al 2012;Yeeles et al 2015) and has been implicated in the separation of the Mcm2-7 double hexamer (Quan et al 2015). It is possible that the double hexamer of the Mcm2-7 complex inhibits DNA unwinding and that Mcm10 activates unwinding by causing double-hexamer separation (Quan et al 2015).…”

mentioning

confidence: 99%

“…In contrast, Mcm10 is required for initial DNA unwinding at origins of replication (Kanke et al 2012;van Deursen et al 2012;Watase et al 2012;Yeeles et al 2015) and has been implicated in the separation of the Mcm2-7 double hexamer (Quan et al 2015). It is possible that the double hexamer of the Mcm2-7 complex inhibits DNA unwinding and that Mcm10 activates unwinding by causing double-hexamer separation (Quan et al 2015). Alternatively, Mcm10 could facilitate extrusion of ssDNA from the Mcm2-7 central channel, enabling the transition from Mcm2-7 encircling dsDNA to ssDNA (Costa et al 2014 (Homesley et al 2000;Apger et al 2010;Lee et al 2010).…”

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confidence: 99%

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Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase

2017

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Activation of the Mcm2-7 replicative DNA helicase is the committed step in eukaryotic DNA replication initiation. Although Mcm2-7 activation requires binding of the helicase-activating proteins Cdc45 and GINS (forming the CMG complex), an additional protein, Mcm10, drives initial origin DNA unwinding by an unknown mechanism. We show that Mcm10 binds a conserved motif located between the oligonucleotide/oligosaccharide fold (OB-fold) and A subdomain of Mcm2. Although buried in the interface between these domains in Mcm2-7 structures, mutations predicted to separate the domains and expose this motif restore growth to conditional-lethal MCM10 mutant cells. We found that, in addition to stimulating initial DNA unwinding, Mcm10 stabilizes Cdc45 and GINS association with Mcm2-7 and stimulates replication elongation in vivo and in vitro. Furthermore, we identified a lethal allele of MCM10 that stimulates initial DNA unwinding but is defective in replication elongation and CMG binding. Our findings expand the roles of Mcm10 during DNA replication and suggest a new model for Mcm10 function as an activator of the CMG complex throughout DNA replication.

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Section: Mcm10 Binds a Conserved Region In The Mcm2 N-terminal Domainsupporting

confidence: 75%

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confidence: 99%

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confidence: 99%

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Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase

2017

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“…The prevalent model from in vitro and in vivo studies in budding and fission yeast is that Mcm10 plays a role late in replication initiation where it is required for unwinding of origin DNA and separation of Mcm2-7 double hexamers 10,26-29 In addition to its involvement in DNA replication initiation, Mcm10 has also been shown to promote genomic integrity in human cells, as lack of Mcm10 leads to accumulation of DNA damage and cell cycle arrest 22,30-32 Budding yeast Mcm10 performs some of its genome protection functions by interactions with 9-1-1 checkpoint clamp and other factors implicated in double strand break repair 33,34 …”

Section: Introductionmentioning

confidence: 99%

Xenopus Mcm10 is a CDK-substrate required for replication fork stability

et al. 2016

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During S phase, following activation of the S phase CDKs and the DBF4-dependent kinases (DDK), double hexamers of Mcm2-7 at licensed replication origins are activated to form the core replicative helicase. Mcm10 is one of several proteins that have been implicated from work in yeasts to play a role in forming a mature replisome during the initiation process. Mcm10 has also been proposed to play a role in promoting replisome stability after initiation has taken place. The role of Mcm10 is particularly unclear in metazoans, where conflicting data has been presented. Here, we investigate the role and regulation of Mcm10 in Xenopus egg extracts. We show that Xenopus Mcm10 is recruited to chromatin late in the process of replication initiation and this requires prior action of DDKs and CDKs. We also provide evidence that Mcm10 is a CDK substrate but does not need to be phosphorylated in order to associate with chromatin. We show that in extracts depleted of more than 99% of Mcm10, the bulk of DNA replication still occurs, suggesting that Mcm10 is not required for the process of replication initiation. However, in extracts depleted of Mcm10, the replication fork elongation rate is reduced. Furthermore, the absence of Mcm10 or its phosphorylation by CDK results in instability of replisome proteins on DNA, which is particularly important under conditions of replication stress.

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“…Crucially, during helicase activation, the MCM2-7 DH becomes extensively reorganized: Cdc45 and GINS bind to MCM2-7; the complex is split into two individual hexamers, potentially involving Mcm10 (Quan et al 2015); one DNA strand becomes extruded from each hexamer; and this results in the two CMG complexes encircling ssDNA (Costa et al 2014;Georgescu et al 2017). Importantly, the mechanisms that lead to this MCM2-7 reorganization during initiation of DNA replication are largely unknown.…”

Section: Initiation Of Dna Replicationmentioning

confidence: 99%

From structure to mechanism—understanding initiation of DNA replication

et al. 2017

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DNA replication results in the doubling of the genome prior to cell division. This process requires the assembly of 50 or more protein factors into a replication fork. Here, we review recent structural and biochemical insights that start to explain how specific proteins recognize DNA replication origins, load the replicative helicase on DNA, unwind DNA, synthesize new DNA strands, and reassemble chromatin. We focus on the minichromosome maintenance (MCM2-7) proteins, which form the core of the eukaryotic replication fork, as this complex undergoes major structural rearrangements in order to engage with DNA, regulate its DNA-unwinding activity, and maintain genome stability.

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Cell-Cycle-Regulated Interaction between Mcm10 and Double Hexameric Mcm2-7 Is Required for Helicase Splitting and Activation during S Phase

Cited by 57 publications

References 45 publications

Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase

Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase

Xenopus Mcm10 is a CDK-substrate required for replication fork stability

From structure to mechanism—understanding initiation of DNA replication

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