Receptor- and phorbol-ester-mediated redistribution of protein kinase C in human platelets. Evidence that aggregation promotes degradation of protein kinase C.
Translocation of Ca2+/phospholipid-dependent protein kinase (PKC) activity from cytosolic to membrane fractions was assessed in washed human platelet suspensions. Phorbol myristate acetate (PMA) induced a rapid loss of PKC activity from the cytosolic compartment in stirred platelets, which was not accompanied by measurable increases in membrane-associated activity, but was paralleled by a decrease in total cellular enzyme activity (cytosol plus membrane). When platelet aggregation was prevented by not stirring, (i) cytosolic activity was decreased by PMA, (ii) significant and maintained (1-15 min with PMA) increases in membrane-bound PKC were detected, and (iii) the decline in total enzyme activity was markedly slower. In stirred platelets, total and specific inhibition of PMA-induced aggregation by a fibrinogen-derived peptide (RGDS, i.e. Arg-Gly-Asp-Ser) promoted maximal increases in membrane-associated PKC in the presence of PMA and completely prevented the loss in cellular activity. Thrombin and collagen both induced a decrease in cytosolic PKC and a loss of total activity, but a significant rise in membrane activity was seen only with collagen; ADP had no detectable effect on enzyme distribution. These results demonstrate an agonist-induced redistribution of PKC and indicate that platelet aggregation may play an important role in the proteolysis, and hence persistence, of membrane-associated PKC. This observation has implications for the potency and duration of PKC-mediated responses induced by agonists and exogenous PKC activators.
Wheeler-Jones
C P
CP
Thrombosis Research Unit, King's College School of Medicine and Dentistry, Rayne Institute, London, U.K.
Patel
Y
Y
Kakkar
V V
VV
Krishnamurthi
S
S
D000244:Adenosine Diphosphate / Q000494:pharmacology; D001792:Blood Platelets / Q000201:enzymology*; D002451:Cell Compartmentation / Q000187:drug effects; D002462:Cell Membrane / Q000201:enzymology; D003094:Collagen / Q000494:pharmacology; D003600:Cytosol / Q000201:enzymology; D006801:Humans; D066298:In Vitro Techniques; D009842:Oligopeptides / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013997:Time Factors
D009842:Oligopeptides; D000244:Adenosine Diphosphate; D003094:Collagen; D011493:Protein Kinase C; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate