Clustering of integrin alpha(IIb)-beta(3) differently regulates tyrosine phosphorylation of pp72(syk), PLC gamma 2 and pp125(FAK) in Concanavalin A-stimulated platelets

Tyrosine phosphorylation of the non-receptor tyrosine kinases pp72(syk) and pp125(FAK) and of the gamma 2 isoform of phospholipase C (PLC gamma 2) in human platelets stimulated with the lectin Concanavalin A was investigated. Concanavalin A induced the rapid tyrosine phosphorylation of pp72(syk) and PLC gamma 2 with a similar kinetics, while tyrosine phosphorylation of pp1 25(FAK) occurred in a later phase of platelet activation. When compared wit h other platelet agonists, Concanavalin A revealed to be at least as potent as collagen in inducing tyrosine phosphorylation of PLC gamma 2 and pp125( FAK), while tyrosine phosphorylation of pp72(syk) induced by the lectin was much stronger than that induced by thrombin or collagen. Concanavalin A-in duced tyrosine phosphorylation of pp72(syk), PLC gamma 2 and pp125(FAK) was not dependent on platelet aggregation as it occurred normally even in the absence of sample stirring and when fibrinogen binding to integrin alpha(II b)-beta(3) was inhibited by the peptide RGDS. Tyrosine phosphorylation of p p72(syk), PLC gamma 2 and pp125(FAK) required the binding of the lectin to the platelet surface, but was not observed in platelets treated with succin yl-Concanavalin A, a derivative of the lectin that interacts with the same receptors but does not promote clustering of membrane glycoproteins. Moreov er the aggregation-independent tyrosine phosphorylation of pp125(FAK) and p p72(syk) induced by Concanavalin A required the expression of integrin alph a(IIb)-beta(3) on the platelet surface as it was strongly inhibited in plat elets from patients affected by Glanzmann thrombasthenia. By contrast, tyro sine phosphorylation of PLC gamma 2 occurred normally also in thrombastheni c platelets stimulated with Concanavalin A. These results demonstrate that, even in the absence of aggregation, the clustering of integrin alpha(IIb)- beta(3) induced by Concanavalin A on the platelet surface directly promotes tyrosine phosphoryIation of pp72(syk) and pp125(FAK) and provide further e vidence that the oligomerization of the fibrinogen receptor promoted by its natural ligand during platelet aggregation may be responsible for the tyro sine phosphorylation of these proteins induced by physiological agonists.