title: Inhibition of protein kinase C by staurosporine promotes elevated accumulations of inositol trisphosphates and tetrakisphosphate in human platelets exposed to thrombin.
abstract: We have examined regulation by protein kinase C (Ca2+/phospholipid-dependent enzyme) of thrombin-induced inositol polyphosphate accumulation in human platelets. When platelets are exposed to thrombin for 10 s, the protein kinase C inhibitor staurosporine causes inositol phosphate elevations over control values of 2.7-fold (inositol 1,4,5-trisphosphate (Ins(1,4,5)P3], 1.9-fold (inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4], and 1.2-fold (inositol 1,3,4-trisphosphate). In the same period, phosphatidic acid and diacylglycerol are unaffected. The myosin light chain kinase inhibitor ML-7 has no effect on inositol phosphate accumulations. Staurosporine does not inhibit Ins(1,4,5)P3 3-kinase and 5-phosphomonoesterase activities in saponin-permeabilized platelets incubated with exogenous Ins(1,4,5)P3 unless the platelets have been exposed to thrombin and protein kinase C is consequently activated. The protein kinase C agonist beta-phorbol 12,13-dibutyrate increases the Vmax of the 3-kinase 1.8-fold, with little effect on Km. Our results provide strong evidence for a role for protein kinase C in regulating inositol phosphate levels in thrombin-activated platelets. We propose that endogenously activated protein kinase C removes Ins(1,4,5)P3 by stimulating both 5-phosphomonoesterase and Ins(1,4,5)P3 3-kinase. Initial activation of phospholipase C does not appear to be affected by such protein kinase C. Inhibition of protein kinase C by staurosporine decreases 5-phosphomonoesterase activity. The resulting elevated Ins(1,4,5)P3, as substrate for Ins(1,4,5)P3 3-kinase, promotes production of Ins(1,3,4,5)P4, which also may accumulate through decreased 5-phosphomonoesterase activity and elevated Ca2+ levels. These factors apparently counteract the inhibitory effect on 3-kinase, yielding a net increase in Ins(1,3,4,5)P4.
authors.affiliation: Department of Biochemistry, University of Vermont College of Medicine, Burlington 05405.
authors.forename: W G
authors.initials: WG
authors.lastname: King
authors.forename: S E
authors.initials: SE
authors.lastname: Rittenhouse
mesh_terms: D000470:Alkaloids / Q000494:pharmacology*; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000097:blood; D004075:Diglycerides / Q000097:blood; D006801:Humans; D066298:In Vitro Techniques; D007295:Inositol Phosphates / Q000097:blood*; D009218:Myosins / Q000378:metabolism; D010712:Phosphatidic Acids / Q000097:blood; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D019311:Staurosporine; D013403:Sugar Phosphates / Q000097:blood*; D013917:Thrombin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D000470:Alkaloids; D004075:Diglycerides; D007295:Inositol Phosphates; D010712:Phosphatidic Acids; D013403:Sugar Phosphates; D011493:Protein Kinase C; D010738:Type C Phospholipases; D013917:Thrombin; D009218:Myosins; D019311:Staurosporine; D002118:Calcium