Okadaic acid, a serine/threonine phosphatase inhibitor, induces tyrosine dephosphorylation/inactivation of protein kinase FA/GSK-3 alpha in A431 cells.
The signal transduction mechanism of protein kinase FA/GSK-3 alpha by tyrosine phosphorylation in A431 cells was investigated. Kinase FA/GSK-3 alpha was found to exist in a highly tyrosine-phosphorylated/activated state in resting cells but could be tyrosine-dephosphorylated and inactivated down to less than 15% of control values in a concentration-dependent manner by 50-400 nM okadaic acid (a specific inhibitor of protein phosphatase types 1 and 2A), as demonstrated by metabolic 32P labeling the cells, followed by immunoprecipitation and two-dimensional phosphoamino acid analysis and by immunodetection in an anti-kinase FA/GSK-3 alpha immunoprecipitate kinase assay. Taken together, the results provide initial evidence that serine/threonine phosphatase(s) may play a role involved in the modulation of kinase FA/GSK-3 alpha activity in cells, suggesting an involvement of serine/threonine dephosphorylation in the modulation of tyrosine phosphorylation and activation of protein kinase FA/GSK-3 alpha, representing a new mode of signal transduction pathway for the regulation of this multisubstrate protein kinase in cells.
Yu
J S
JS
Institute of Biomedical Sciences, National Tsing Hua University, Hsinchu, China.
Yang
S D
SD
D000470:Alkaloids / Q000494:pharmacology; D000595:Amino Acid Sequence; D000906:Antibodies; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004988:Ethers, Cyclic / Q000494:pharmacology*; D038362:Glycogen Synthase Kinase 3; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D019319:Okadaic Acid; D009842:Oligopeptides / Q000737:chemistry / Q000276:immunology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D019000:Phosphotyrosine; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019311:Staurosporine; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D000470:Alkaloids; D000906:Antibodies; D004988:Ethers, Cyclic; D009842:Oligopeptides; D019319:Okadaic Acid; D019000:Phosphotyrosine; D014443:Tyrosine; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D010749:Phosphoprotein Phosphatases; D019311:Staurosporine