Fate of immunoprecipitable protein kinase C in GH3 cells treated with phorbol 12-myristate 13-acetate.
The effect of phorbol 12-myristate 13-acetate (PMA) on protein kinase C was studied by metabolically labeling GH3 cells with [35S]methionine and using a polyclonal antibody raised against rat brain protein kinase C to immunoprecipitate the enzyme. PMA accelerates the loss of immunologically reactive protein kinase C from the cells in a time- and dose-dependent manner. The half-life of the enzyme in cells treated with 400 nM PMA was 2 h whereas in control cells 60-70% of the enzyme was still detectable after 24 h. The concentration of PMA required to reduce cellular protein kinase C 50% after 24 h was 130 nM. PMA also induced the translocation of [35S]Met-labeled protein kinase C from the cytosol to the membranes in a concentration-dependent manner. Less protein kinase C was translocated to membranes when cells were treated with 20 nM PMA than when they were exposed to 400 nM PMA. In the latter case, most of the labeled protein kinase C became membrane-associated. Maximal translocation was evident after 15 min of incubation with either concentration of PMA and was followed by degradation of the membrane-associated enzyme. The rate of degradation of membrane-associated protein kinase C was the same with both concentrations of PMA. In cells treated with 20 nM PMA, disappearance of [35S]Met-labeled protein kinase C from the cytosolic fraction occurred in two phases, a rapid decrease characteristic of the membrane-associated enzyme, followed by a slower loss similar to that seen in control cells. The results indicate that turnover of protein kinase C is enhanced by membrane association.
D000818:Animals; D001921:Brain / Q000201:enzymology; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D006207:Half-Life; D007106:Immune Sera; D007163:Immunosorbent Techniques; D008715:Methionine / Q000378:metabolism; D010704:Phorbols / Q000494:pharmacology*; D010911:Pituitary Neoplasms / Q000201:enzymology*; D011493:Protein Kinase C / Q000276:immunology / Q000378:metabolism*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013997:Time Factors
D007106:Immune Sera; D010704:Phorbols; D008715:Methionine; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate