Role of the cdc25C phosphatase in G2 arrest induced by nitrogen mustard.
G2 arrest induced by nitrogen mustard in human lymphoma CA46 cells is associated with a failure to activate hyperphosphorylated cdc2/cyclin B1 complexes. We investigated the possibility that this might be due to a suppression of cdc25C phosphatase activity. cdc25C from interphase cells migrated as a 54- to 57-kDa doublet in SDS gels and exhibited basal phosphatase activity. cdc25C from mitotic cells migrated as a 66-kDa hyperphosphorylated species and exhibited elevated phosphatase activity. cdc25C hyperphosphorylation and activation were mediated by cdc2, supporting the view of a cdc2-cdc25C autocatalytic feedback loop. Immunofluorescence and cell fractionation studies suggested cdc2-cdc25C interaction occurred within the cytoplasm. Cells arrested in G2 phase following nitrogen mustard treatment or cells arrested in S phase with aphidicolin failed to dephosphorylate and activate cdc2, and this correlated with failure to convert cdc25C into the most active hyperphosphorylated species. Our findings suggest that checkpoints guarding against mitotic entry in the presence of unreplicated or damaged DNA suppress formation of the cdc2-cdc25C autocatalytic feedback loop that normally brings about rapid activation of cdc2.
O'Connor
P M
PM
Division of Cancer Treatment, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Ferris
D K
DK
Hoffmann
I
I
Jackman
J
J
Draetta
G
G
Kohn
K W
KW
D016590:Aphidicolin / Q000494:pharmacology; D002051:Burkitt Lymphoma; D016203:CDC2 Protein Kinase / Q000378:metabolism; D018797:Cell Cycle Proteins; D002460:Cell Line; D016195:G2 Phase / Q000187:drug effects / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007399:Interphase; D007700:Kinetics; D008466:Mechlorethamine / Q000494:pharmacology*; D015739:Nocodazole / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured; D020687:cdc25 Phosphatases
D018797:Cell Cycle Proteins; D011506:Proteins; D016590:Aphidicolin; D008466:Mechlorethamine; D016203:CDC2 Protein Kinase; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; D015739:Nocodazole