Inhibition of p34cdc2 kinase activity by etoposide or irradiation as a mechanism of G2 arrest in Chinese hamster ovary cells.
The mammalian homologue of the yeast cdc2 gene product, p34cdc2, is a cell cycle-regulated protein essential for mitosis. We have used polyclonal antisera raised against a peptide corresponding to the carboxyl terminus of the sequence of human cdc2 to study p34cdc2 in Chinese hamster ovary (CHO) cells. Major bands are immunoprecipitated at a molecular weight of 34,000, although not in the presence of competing antigenic peptide. p34cdc2 is coimmunoprecipitated with proteins of molecular weights of 52,000 and 57,000. Immunoprecipitates express histone H1 kinase activity which varies throughout the cell cycle, maximal activity being observed in G2-M. The activity of the p34cdc2 kinase varies according to its association with the Mr 52,000 and 57,000 proteins and according to their phosphorylation state. Treatment of either asynchronous CHO cells or an enriched G2 population with the antitumor agent, etoposide, results in rapid inhibition of immunoprecipitated p34cdc2 kinase activity, which is not due to a direct effect of drug upon the enzyme. p34cdc2 kinase activity recovers as cells arrest in G2 and a second etoposide treatment further inhibits p34cdc2 kinase activity and prolongs G2 arrest. Exposure of asynchronous CHO cells to gamma-irradiation also inhibits p34cdc2 kinase activity within 1 h. Again this activity recovers as cells accumulate in G2. These results suggest that DNA damage in CHO cells elicits a response which results in inhibition of p34cdc2 kinase activity and, consequently, G2 arrest.
Graham Brown Cancer Center, University of Louisville, Kentucky 40292.
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D000818:Animals; D016203:CDC2 Protein Kinase; D002453:Cell Cycle / Q000187:drug effects / Q000528:radiation effects; D002460:Cell Line; D006224:Cricetinae; D004247:DNA / Q000528:radiation effects; D004249:DNA Damage; D005047:Etoposide / Q000494:pharmacology; D008970:Molecular Weight; D010750:Phosphoproteins / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000528:radiation effects; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000032:analysis / Q000378:metabolism / Q000528:radiation effects; D013997:Time Factors
D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; D005047:Etoposide; D004247:DNA; D011494:Protein Kinases; D016203:CDC2 Protein Kinase