Biphasic activation of two mitogen-activated protein kinases during the cell cycle in mammalian cells.
We studied mitogen-activated protein kinase (MAPK) activities during the cell cycle of Chinese hamster ovary (CHO) cells using site-specific antibodies against extracellular signal-regulated kinase-1, a 44-kDa MAPK (Boulton, T.G., Yancopoulos, G.D., Gregory, J.S., Slauer, C., Moomaw, C., Hsu, J., and Cobb, M.H. (1990) Science 249, 64-67). These antibodies detected two distinct MAPKs (44- and 42-kDa MAPKs) in CHO cells. CHO cells were arrested at metaphase in the M phase by treatment with nocodazole, and activities of MAPKs were analyzed at specific time points after release from arrest. Immune complex kinase assay and renaturation and phosphorylation assay in substrate-containing gel revealed that both 44- and 42-kDa MAPKs had activities in the G1 through S and G2/M phases and were activated biphasically, in the G1 phase and around the M phase. MAPKs were inactivated in metaphase-arrested cells. The amount of MAPKs did not change significantly in the cell cycle. In the G1, S, and G2/M phases, MAPKs were phosphorylated on both tyrosine and threonine residues and dephosphorylated in metaphase-arrested cells. Our data suggest that MAPKs may play some role in the cell cycle other than G0/G1 transition.
Tamemoto
H
H
Institute for Diabetes Care and Research, Asahi Life Foundation, Tokyo, Japan.
Kadowaki
T
T
Tobe
K
K
Ueki
K
K
Izumi
T
T
Chatani
Y
Y
Kohno
M
M
Kasuga
M
M
Yazaki
Y
Y
Akanuma
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D016203:CDC2 Protein Kinase / Q000378:metabolism; D016466:CHO Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002453:Cell Cycle / Q000187:drug effects; D006224:Cricetinae; D004789:Enzyme Activation; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D008969:Molecular Sequence Data; D015739:Nocodazole / Q000494:pharmacology; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D013912:Threonine; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D016203:CDC2 Protein Kinase; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D015739:Nocodazole; D013936:Thymidine