Role of inhibitory CDC2 phosphorylation in radiation-induced G2 arrest in human cells.
The activity of the mitosis-promoting kinase CDC2-cyclin B is normally suppressed in S phase and G2 by inhibitory phosphorylation at Thr14 and Tyr15. This work explores the possibility that these phosphorylations are responsible for the G2 arrest that occurs in human cells after DNA damage. HeLa cell lines were established in which CDC2AF, a mutant that cannot be phosphorylated at Thr14 and Tyr15, was expressed from a tetracycline-repressible promoter. Expression of CDC2AF did not induce mitotic events in cells arrested at the beginning of S phase with DNA synthesis inhibitors, but induced low levels of premature chromatin condensation in cells progressing through S phase and G2. Expression of CDC2AF greatly reduced the G2 delay that resulted when cells were X-irradiated in S phase. However, a significant G2 delay was still observed and was accompanied by high CDC2-associated kinase activity. Expression of wild-type CDC2, or the related kinase CDK2AF, had no effect on the radiation-induced delay. Thus, inhibitory phosphorylation of CDC2, as well as additional undefined mechanisms, delay mitosis after DNA damage.
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Department of Physiology, University of California, San Francisco 94143-0444, USA.
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Morgan
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D016203:CDC2 Protein Kinase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D042846:CDC2-CDC28 Kinases; D019926:Cyclin B; D056744:Cyclin B1; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000096:biosynthesis; D016213:Cyclins / Q000378:metabolism; D004249:DNA Damage / Q000502:physiology*; D004794:Enzyme Repression / Q000187:drug effects; D016195:G2 Phase / Q000502:physiology* / Q000528:radiation effects; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology; D009154:Mutation; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D016196:S Phase / Q000502:physiology / Q000528:radiation effects; D013752:Tetracycline / Q000494:pharmacology
C534699:CCNB1 protein, human; D019926:Cyclin B; D056744:Cyclin B1; D016213:Cyclins; D011500:Protein Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D013752:Tetracycline