p34(Cdc2) kinase activity is excluded from the nucleus during the radiation-induced G(2) arrest in HeLa cells.
The progression of cells from G(2) into mitosis is blocked by exposure to DNA-damaging agents such as ionizing radiation. This G(2) delay is associated with reduced cyclin B1-specific associated histone H1 kinase activity, increased inhibitory phosphorylation of p34(Cdc2), and depressed cyclin B1 levels in HeLa cells. Induction of cyclin B1 or expression of Cdc2AF, a mutant p34(Cdc2) that lacks the sites of inhibitory phosphorylation, only partially reverses the radiation-associated G(2) delay, although both maneuvers rapidly result in increased histone H1 kinase activity. To account for the persistent G(2) delay in the face of active p34(Cdc2) kinase, we determined the location of the kinase activity. Although p34(Cdc2) was active in the cytoplasm, the nuclear p34(Cdc2) was inactive. Irradiation led to nuclear accumulation of the inactive tyrosine-phosphorylated form of p34(Cdc2), whereas the active form was seen in the cytoplasm. At later times when cells had resumed cell cycle progression, nuclear kinase activity was detectable. These results give evidence of segregation of cytoplasmic and nuclear kinase activity after DNA damage that has the effect of enhancing checkpoint control. Shielding the nucleus from the potentially deleterious effects of kinase activity after DNA damage may help irradiated human cancer cells respond to irradiation.
Kao
G D
GD
Department of Radiation Oncology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
McKenna
W G
WG
Muschel
R J
RJ
D016203:CDC2 Protein Kinase / Q000378:metabolism*; D002467:Cell Nucleus / Q000201:enzymology* / Q000378:metabolism / Q000528:radiation effects; D019926:Cyclin B / Q000378:metabolism*; D056744:Cyclin B1; D003593:Cytoplasm / Q000201:enzymology / Q000378:metabolism; D004249:DNA Damage; D003907:Dexamethasone / Q000494:pharmacology; D004307:Dose-Response Relationship, Radiation; D016195:G2 Phase / Q000528:radiation effects*; D015971:Gene Expression Regulation, Enzymologic / Q000528:radiation effects*; D006367:HeLa Cells; D006801:Humans; D016200:Maturation-Promoting Factor / Q000378:metabolism; D008938:Mitosis / Q000187:drug effects / Q000528:radiation effects; D010766:Phosphorylation; D013997:Time Factors; D014404:Tubulin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
C534699:CCNB1 protein, human; D019926:Cyclin B; D056744:Cyclin B1; D014404:Tubulin; D014443:Tyrosine; D003907:Dexamethasone; D016203:CDC2 Protein Kinase; D016200:Maturation-Promoting Factor