Cdk1-mediated phosphorylation of Cdc7 suppresses DNA re-replication.
To maintain genetic stability, the entire mammalian genome must replicate only once per cell cycle. This is largely achieved by strictly regulating the stepwise formation of the pre-replication complex (pre-RC), followed by the activation of individual origins of DNA replication by Cdc7/Dbf4 kinase. However, the mechanism how Cdc7 itself is regulated in the context of cell cycle progression is poorly understood. Here we report that Cdc7 is phosphorylated by a Cdk1-dependent manner during prometaphase on multiple sites, resulting in its dissociation from origins. In contrast, Dbf4 is not removed from origins in prometaphase, nor is it degraded as cells exit mitosis. Our data thus demonstrates that constitutive phosphorylation of Cdc7 at Cdk1 recognition sites, but not the regulation of Dbf4, prevents the initiation of DNA replication in normally cycling cells and under conditions that promote re-replication in G2/M. As cells exit mitosis, PP1α associates with and dephosphorylates Cdc7. Together, our data support a model where Cdc7 (de)phosphorylation is the molecular switch for the activation and inactivation of DNA replication in mitosis, directly connecting Cdc7 and PP1α/Cdk1 to the regulation of once-per-cell cycle DNA replication in mammalian cells.
a Tumour Biology Group, Advanced Medical Research Institute of Canada, Health Sciences North , Sudbury , Ontario , Canada.
James
J
Knockleby
a Tumour Biology Group, Advanced Medical Research Institute of Canada, Health Sciences North , Sudbury , Ontario , Canada.
Byung Ju
BJ
Kim
a Tumour Biology Group, Advanced Medical Research Institute of Canada, Health Sciences North , Sudbury , Ontario , Canada.
Avani
A
Mehta
a Tumour Biology Group, Advanced Medical Research Institute of Canada, Health Sciences North , Sudbury , Ontario , Canada.|b Departments of Medicine , the Faculty of Medicine, the University of Ottawa , Ottawa , Ontario , Canada.|c Northern Ontario School of Medicine , Sudbury , Ontario , Canada.
Hoyun
H
Lee
D016203:CDC2 Protein Kinase; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000235:genetics* / Q000378:metabolism; D004261:DNA Replication; D059565:G2 Phase Cell Cycle Checkpoints; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008938:Mitosis; D010766:Phosphorylation; D054645:Protein Phosphatase 1 / Q000235:genetics* / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D000097573:Red Fluorescent Protein
D016203:CDC2 Protein Kinase; D018797:Cell Cycle Proteins; D018844:Cyclin-Dependent Kinases; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D054645:Protein Phosphatase 1; D017346:Protein Serine-Threonine Kinases; D000097573:Red Fluorescent Protein; C485678:DBF4 protein, human; C485184:enhanced green fluorescent protein; C068619:CDC7 protein, human; C528899:CDK1 protein, human
Cdc7; DDK; DNA replication; cell cycle; phosphorylation