title: Direct regulation of Treslin by cyclin-dependent kinase is essential for the onset of DNA replication.
abstract: Treslin, a TopBP1-interacting protein, is necessary for deoxyribonucleic acid (DNA) replication in vertebrates. Association between Treslin and TopBP1 requires cyclin-dependent kinase (Cdk) activity in Xenopus laevis egg extracts. We investigated the mechanism and functional importance of Cdk for this interaction using both X. laevis egg extracts and human cells. We found that Treslin also associated with TopBP1 in a Cdk-regulated manner in human cells and that Treslin was phosphorylated within a conserved Cdk consensus target sequence (on S976 in X. laevis and S1000 in humans). Recombinant human Cdk2-cyclin E also phosphorylated this residue of Treslin in vitro very effectively. Moreover, a mutant of Treslin that cannot undergo phosphorylation on this site showed significantly diminished binding to TopBP1. Finally, human cells harboring this mutant were severely deficient in DNA replication. Collectively, these results indicate that Cdk-mediated phosphorylation of Treslin during S phase is necessary for both its effective association with TopBP1 and its ability to promote DNA replication in human cells.
authors.affiliation: Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.
authors.forename: Akiko
authors.initials: A
authors.lastname: Kumagai
authors.forename: Anna
authors.initials: A
authors.lastname: Shevchenko
authors.forename: Andrej
authors.initials: A
authors.lastname: Shevchenko
authors.forename: William G
authors.initials: WG
authors.lastname: Dunphy
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D019927:Cyclin E / Q000235:genetics / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2 / Q000235:genetics / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016196:S Phase; D016415:Sequence Alignment; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis / Q000235:genetics / Q000378:metabolism
chemical_list: C475820:CCNE1 protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D019927:Cyclin E; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D011994:Recombinant Proteins; C547407:TICRR protein, Xenopus; C547406:TICRR protein, human; C110513:TOPBP1 protein, human; C509993:TopBP1 protein, Xenopus; D029867:Xenopus Proteins; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases