title: Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase.
abstract: 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-lethalMCM10mutant 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 ofMCM10that 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.
authors.affiliation: Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 USA.
authors.forename: Marko
authors.initials: M
authors.lastname: Lõoke
authors.affiliation: Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 USA.|Microbiology Graduate Program, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 USA.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Maloney
authors.affiliation: Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 USA.
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Bell
mesh_terms: D000595:Amino Acid Sequence; D004265:DNA Helicases / Q000737:chemistry / Q000378:metabolism; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D064110:Minichromosome Maintenance Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D018741:Replication Origin; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D061805:Transcription Elongation, Genetic
chemical_list: C102209:CDC45 protein, S cerevisiae; D004268:DNA-Binding Proteins; C076365:MCM10 protein, S cerevisiae; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D004265:DNA Helicases; C069199:MCM2 protein, S cerevisiae; D064110:Minichromosome Maintenance Proteins
keywords: Cdc45/Mcm2–7/GINS; DNA replication fork; S. cerevisiae; cell cycle; reconstituted DNA replication