Thrombin activation of human platelets dissociates a complex containing gelsolin and actin from phosphatidylinositide-specific phospholipase Cgamma1.
We have examined the association of two cytoskeleton proteins, gelsolin and actin, with phosphatidylinositide-specific phospholipase Cgamma1 (PLCgamma1) in resting and thrombin-stimulated human platelets. In unstimulated platelets, gelsolin, actin and PLCgamma1 were immunoprecipitated as a complex by a polyclonal antibody to PLCgamma1. The association of gelsolin and actin was specific for PLCgamma1 because immunoprecipitates of PLCs beta2, beta3, gamma2 and delta1, which are also expressed in human platelets, did not contain detectable gelsolin or actin. Activation with thrombin resulted in platelet aggregation and the dissociation of gelsolin and actin from PLCgamma1. Inhibition of thrombin-induced platelet aggregation blocked the dissociation of gelsolin and actin from PLCgamma1. After stimulation with thrombin, PLCgamma1 activity in immunoprecipitates was increased 2-3-fold. This elevation in PLCgamma1 activity in response to thrombin activation was not observed when platelet aggregation was blocked. Although PLCgamma1 is tyrosine phosphorylated in response to many agonists, we could not detect, by Western analysis with anti-phosphotyrosine antibodies, tyrosine phosphorylation of PLCgamma1 immunoprecipitated from thrombin-stimulated platelets. These results demonstrate that PLCgamma1 is associated with gelsolin and actin in resting platelets, and that thrombin-induced platelet aggregation results in the dissociation of PLCgamma1 from gelsolin and actin, and the stimulation of PLCgamma1 activity.
Baldassare
J J
JJ
Department of Pharmacological and Physiological Science, St. Louis Health Science Center, St. Louis, MO 63110, USA.
Henderson
P A
PA
Tarver
A
A
Fisher
G J
GJ
D000199:Actins / Q000097:blood* / Q000302:isolation & purification / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D018260:Gelsolin / Q000097:blood* / Q000302:isolation & purification / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000097:blood* / Q000302:isolation & purification; D007700:Kinetics; D051966:Phospholipase C gamma; D015539:Platelet Activation; D010974:Platelet Aggregation; D013917:Thrombin / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000097:blood* / Q000302:isolation & purification
D000199:Actins; D018260:Gelsolin; D007527:Isoenzymes; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D013917:Thrombin