Biphasic activation of two mitogen-activated protein kinases during the cell cycle in mammalian cells.
- H Tamemoto,
- T Kadowaki,
- K Tobe,
- K Ueki,
- T Izumi,
- Y Chatani,
- M Kohno,
- M Kasuga,
- Y Yazaki and
- Y Akanuma
Abstract
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.
