title: Phosphorylation of the inhibitory guanine-nucleotide-binding protein as a possible mechanism of inhibition by protein kinase C of agonist-induced Ca2+ mobilization in human platelet.
abstract: Increases in the intracellular Ca2+ concentration of human platelets caused by receptor agonists, such as thrombin, 9,11-epithio-11,12-methanothromboxane A2 (STA2), platelet-activating factor (PAF) and arginine-vasopressin, were inhibited by prior addition of 12-O-tetradecanoylphorbol 13-acetate (TPA) in time-dependent and concentration-dependent manners. The inhibitions were mostly reversed by staurosporine, and inhibitor of protein kinase C, added 1 min before TPA. Prior treatment of platelets with thrombin or STA2, the efficacious Ca2+ mobilizer, suppressed the increase in the intracellular Ca2+ concentration of the cells to other agonists, but treatment with less efficacious PAF or vasopressin did not. The heterologous receptor desensitizations were also reversed by staurosporine. The antibody, directed against the carboxy-terminal region of the alpha subunits 1 and 2 of the inhibitory guanine-nucleotide-binding proteins (Gi1 alpha and Gi2 alpha), was raised in rabbit and was used to immunoprecipitate Gi alpha in 32P-labeled platelets. The radioactivity was detected in Gi alpha after incubation of 32P-labeled platelets with TPA, thrombin or STA2, but not in the cells incubated with PAF or vasopressin. The time-dependency or concentration-dependency of TPA-induced phosphorylation of Gi alpha was similar to the dependency of its inhibitory action on agonist-induced Ca2+ mobilization. Thus, strong activation of Ca2+/phospholipid-dependent protein kinase C by phorbol ester or agonists of certain Ca(2+)-mobilizing receptors leads to phosphorylation of the alpha subunit of guanine-nucleotide-binding protein, thereby impairing the coupling of the G protein to receptors as a feedback regulatory component of the receptor-triggered intracellular Ca(2+)-mobilizing system.
authors.affiliation: Department of Physiological Chemistry, Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Yatomi
authors.forename: Y
authors.initials: Y
authors.lastname: Arata
authors.forename: S
authors.initials: S
authors.lastname: Tada
authors.forename: S
authors.initials: S
authors.lastname: Kume
authors.forename: M
authors.initials: M
authors.lastname: Ui
mesh_terms: D000470:Alkaloids / Q000494:pharmacology; D000595:Amino Acid Sequence; D001127:Arginine Vasopressin / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010972:Platelet Activating Factor / Q000494:pharmacology; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013928:Thromboxane A2 / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D000470:Alkaloids; D010972:Platelet Activating Factor; D001127:Arginine Vasopressin; D013928:Thromboxane A2; C037866:STA 2; D011493:Protein Kinase C; D013917:Thrombin; D019204:GTP-Binding Proteins; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium