Role of rap1B and p21ras GTPase-activating protein in the regulation of phospholipase C-gamma 1 in human platelets.
Thrombin activates phospholipase C in human platelets, but the specific isoenzymes activated and the signal pathway used are unknown. Using specific antibodies, we found that phospholipase C-gamma 1 and the p21ras GTPase-activating protein, rasGAP, are present in human platelets. Furthermore, phospholipase C-gamma 1 was detectable, based on enzyme activity and Western blot analysis, in immunoprecipitates of rasGAP, suggesting that these two proteins form tight complexes. The pool of phospholipase C-gamma 1 associated with rasGAP was phosphorylated but not through tyrosine phosphorylation. Although thrombin stimulation had no effect on the level of phosphorylation of phospholipase C-gamma 1 and only slightly increased the tyrosine phosphorylation of rasGAP, the agonist induced the association of rasGAP with rap1B, as indicated by the appearance of rap1B on a Western blot of rasGAP immunoprecipitates. Our results suggest the formation of a signaling complex involving rasGAP, phospholipase C-gamma 1, and rap1B that might be important in the cascade leading to platelet activation.
Torti
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Division of Cell Biology, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Lapetina
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D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology*; D001798:Blood Proteins / Q000302:isolation & purification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000097:blood*; D007700:Kinetics; D019269:Phosphatidylinositol 4,5-Diphosphate; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010716:Phosphatidylinositols / Q000378:metabolism; D010750:Phosphoproteins / Q000097:blood / Q000302:isolation & purification; D010727:Phosphoric Diester Hydrolases / Q000096:biosynthesis / Q000097:blood* / Q000302:isolation & purification; D010766:Phosphorylation; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013917:Thrombin / Q000494:pharmacology; D020668:rap GTP-Binding Proteins; D020703:ras GTPase-Activating Proteins
D001798:Blood Proteins; D020690:GTPase-Activating Proteins; D007527:Isoenzymes; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D010750:Phosphoproteins; D011506:Proteins; D020703:ras GTPase-Activating Proteins; D010727:Phosphoric Diester Hydrolases; D013917:Thrombin; D019204:GTP-Binding Proteins; D020668:rap GTP-Binding Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase