Platelet adhesion and aggregation during hemostasis and thrombosis are usually limited to sites where the integrity of the vessel wall is disrupted. The high concentration of platelet agonists within these sites represents a putative control mechanism for targeting platelet activation. Although much has been learned about the intracellular signaling systems controlling platelet activation, our understanding of the connection between signaling molecules and platelet aggregation remains limited. Tyrosine kinases are important signaling enzymes in cells and are abundant in platelets. Previous reports indicate that binding of glycoprotein IIb-IIIa (GPIIb-IIIa) to fibrinogen can induce the tyrosine phosphorylation of specific substrates. We show that, in turn, protein tyrosine kinase activity is necessary for agonist-induced activation of GPIIb-IIIa. Genistein and the tyrphostin AG-18 are two specific tyrosine kinase inhibitors, and the former has been shown to inhibit platelet aggregation. We use genistein and AG-18 in the present study to demonstrate that aggregation inhibition is due to suppression of GPIIb- IIIa activation. In contrast, genistin, an isoflavone compound related to genistein, and acetylsalicylic acid do not affect the tyrosine kinase- signaling pathway, nor do they inhibit GPIIb-IIIa activation induced by strong agonists. On identifying prominent tyrosine kinase substrates in activated platelets, we confirm that several substrates correspond to proteins associated with the cytoskeleton: the 85-kD subunit of phosphatidylinositol 3-kinase, the SH3-containing and actin-associating p85, pp60(Src), and pp125(FAK). Our data showing that tyrosine kinase activity is required for GPIIb-IIIa activation, together with previous studies indicating that fibrinogen binding to its receptor results in tyrosine phosphorylation of platelet substrates, suggest that a dual regulatory mechanism allows for full platelet response only at sites where both pathways are activated, namely disrupted vessel walls.
- glycoprotein IIb-IIIa
- tyrosine kinases
ASJC Scopus subject areas
- Cardiology and Cardiovascular Medicine