Okadaic acid activates microtubule-associated protein kinase in quiescent fibroblastic cells.
Okadaic acid is a potent and specific inhibitor of protein phosphatases 1 and 2A, and is a strong tumor promoter that is not an activator of protein kinase C. Treatment of quiescent cultures of rat fibroblastic 3Y1 cells with okadaic acid induced marked activation of a kinase activity that phosphorylated microtubule-associated protein (MAP) 2 and myelin basic protein, but not histone or casein, in vitro. This activated kinase eluted at approximately 0.15 M NaCl on a DEAE-cellulose column and its apparent molecular mass was determined to be approximately 40 kDa by gel filtration. Detection of the kinase activity in polyacrylamide gels containing substrate proteins after sodium dodecyl sulfate gel electrophoresis revealed that the okadaic-acid-activated kinase activity resided mainly in two closely related polypeptides with apparent molecular mass approximately 40 kDa. The characteristics of this kinase were indistinguishable from those of the mitogen-activated MAP kinase in the same cells. The okadaic-acid-activated MAP kinase was deactivated by protein phosphatase 2A treatment in vitro. These results suggest that MAP kinase is negatively regulated by protein phosphatases 1 and/or 2A in quiescent cells and therefore can be activated by inhibiting these protein phosphatases. Interestingly, the okadaic-acid-induced activation of MAP kinase was transient and epidermal-growth-factor-induced activation was also transient, even in the presence of okadaic acid. These data may imply that protein phosphatases 1 and 2A are not involved in the deactivation of MAP kinase in cells.
Gotoh
Y
Y
Department of Biophysics and Biochemistry, Faculty of Science, University of Tokyo, Japan.
Nishida
E
E
Sakai
H
H
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002460:Cell Line; D004789:Enzyme Activation; D004988:Ethers, Cyclic / Q000494:pharmacology*; D005347:Fibroblasts / Q000201:enzymology; D007700:Kinetics; D019319:Okadaic Acid; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats
D004988:Ethers, Cyclic; D019319:Okadaic Acid; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases