Co-activation of protein kinase C and NADPH oxidase in the plasma membrane of neutrophil cytoplasts.
Enucleated, granule-free neutrophil cytoplasts, which in hypotonic media fully release cytosolic components and generate ghosts, have been used to study the cell localization of protein kinase C (PK-C). Treatment of cytoplasts with phorbol myristate acetate, a potent activator of neutrophil functions, triggers translocation of PK-C from the cytosol to the plasma membrane, with an activity recovery of 83 +/- 16%. In the ghost fraction, PK-C catalyzes the phosphorylation of polypeptides with an apparent mol. wt. of 115K, 89K, 79K, 62K, 47K and 19K. From the plasma membrane PK-C can be extracted in an active form by Triton X-100 but not by EGTA. Translocation of PK-C is already evident at 5 sec and plateaus at about 50 sec. Activation of plasmalemmal, O-2 generating NADPH oxidase by the phorbol ester is delayed by about 20 sec with respect to the activation of PK-C. Dose/response experiments show that the pattern of activation of O-2 generation by cytoplasts strictly superimposes with the pattern of PK-C translocation.
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006657:Histones / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D019255:NADPH Oxidases; D009504:Neutrophils / Q000201:enzymology*; D011493:Protein Kinase C / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D006657:Histones; D013481:Superoxides; D000255:Adenosine Triphosphate; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate