title: Chemical genetic analyses of quantitative changes in Cdk1 activity during the human cell cycle.
abstract: Cyclin-dependent kinase 1 (Cdk1) controls cell proliferation and is inhibited by promising anticancer agents, but its mode of action and the consequences of its inhibition are incompletely understood. Cdk1 promotes S- and M-phases during the cell-cycle but also suppresses endoreduplication, which is associated with polyploidy and genome instability. The complexity of Cdk1 regulation has made it difficult to determine whether these different roles require different thresholds of kinase activity and whether the surge of activity as inhibitory phosphates are removed at mitotic onset is essential for cell proliferation. Here, we have used chemical genetics in a human cell line to address these issues. We rescued cells lethally depleted of endogenous Cdk1 with an exogenous Cdk1 conferring sensitivity to one ATP analogue inhibitor (1NMPP1) and resistance to another (RO3306). At no 1NMPP1 concentration was mitosis in rescued clones prevented without also inducing endoreduplication, suggesting that these two key roles for Cdk1 are not simply controlled by different Cdk1 activity thresholds. We also rescued RO3306-resistant clones using exogenous Cdk1 without inhibitory phosphorylation sites, indicating that the mitotic surge of Cdk1 activity is dispensable for cell proliferation. These results suggest that the basic mammalian cycle requires at least some qualitative changes in Cdk1 activity and that quantitative increases in activity need not be rapid. Furthermore, the viability of cells that are unable to undergo rapid Cdk1 activation, and the strong association between endoreduplication and impaired proliferation, may place restrictions on the therapeutic use of a Cdk1 inhibitors.
authors.affiliation: Gene Targeting Group, Centre for Haematology, Imperial College Faculty of Medicine, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
authors.forename: Polly
authors.initials: P
authors.lastname: Gravells
authors.forename: Kazunori
authors.initials: K
authors.lastname: Tomita
authors.forename: Alexander
authors.initials: A
authors.lastname: Booth
authors.forename: Joshua
authors.initials: J
authors.lastname: Poznansky
authors.forename: Andrew C G
authors.initials: AC
authors.lastname: Porter
mesh_terms: D000818:Animals; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D002453:Cell Cycle / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002455:Cell Division; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D016195:G2 Phase; D006801:Humans; D008938:Mitosis / Q000187:drug effects; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D014981:Xenopus; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C582212:1NMPP1 compound; D018797:Cell Cycle Proteins; D011720:Pyrazoles; D011743:Pyrimidines; D029867:Xenopus Proteins; D011494:Protein Kinases; C070577:CDK1 protein, Xenopus; D016203:CDC2 Protein Kinase