title: Xenopus Cdc7 executes its essential function early in S phase and is counteracted by checkpoint-regulated protein phosphatase 1.
abstract: The initiation of DNA replication requires two protein kinases: cyclin-dependent kinase (Cdk) and Cdc7. Although S phase Cdk activity has been intensively studied, relatively little is known about how Cdc7 regulates progression through S phase. We have used a Cdc7 inhibitor, PHA-767491, to dissect the role of Cdc7 in Xenopus egg extracts. We show that hyperphosphorylation of mini-chromosome maintenance (MCM) proteins by Cdc7 is required for the initiation, but not for the elongation, of replication forks. Unlike Cdks, we demonstrate that Cdc7 executes its essential functions by phosphorylating MCM proteins at virtually all replication origins early in S phase and is not limiting for progression through the Xenopus replication timing programme. We demonstrate that protein phosphatase 1 (PP1) is recruited to chromatin and rapidly reverses Cdc7-mediated MCM hyperphosphorylation. Checkpoint kinases induced by DNA damage or replication inhibition promote the association of PP1 with chromatin and increase the rate of MCM dephosphorylation, thereby counteracting the previously completed Cdc7 functions and inhibiting replication initiation. This novel mechanism for regulating Cdc7 function provides an explanation for previous contradictory results concerning the control of Cdc7 by checkpoint kinases and has implications for the use of Cdc7 inhibitors as anti-cancer agents.
authors.affiliation: Centre for Gene Regulation and Expression, College of Life Sciences, University of Dundee, Dow St., Dundee DD1 5EH, UK.
authors.forename: Wei Theng
authors.initials: WT
authors.lastname: Poh
authors.forename: Gaganmeet Singh
authors.initials: GS
authors.lastname: Chadha
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Gillespie
authors.forename: Philipp
authors.initials: P
authors.lastname: Kaldis
authors.forename: J Julian
authors.initials: JJ
authors.lastname: Blow
mesh_terms: D000818:Animals; D016466:CHO Cells; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004261:DNA Replication / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005047:Etoposide / Q000494:pharmacology; D010063:Ovum / Q000201:enzymology / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010881:Piperidones / Q000494:pharmacology; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011758:Pyrroles / Q000494:pharmacology; D059807:S Phase Cell Cycle Checkpoints / Q000187:drug effects; D014981:Xenopus / Q000254:growth & development / Q000378:metabolism; D029867:Xenopus Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D004791:Enzyme Inhibitors; C531237:PHA 767491; D010881:Piperidones; D011758:Pyrroles; D029867:Xenopus Proteins; D005047:Etoposide; C489405:CDC7 protein, Xenopus; D017346:Protein Serine-Threonine Kinases; D054645:Protein Phosphatase 1
keywords: Cdc7; DNA replication; PHA-767491; PP1; Xenopus