Distinct contributions of glycoprotein VI and alpha(2)beta(1) integrin to the induction of platelet protein tyrosine phosphorylation and aggregation

Platelet activation by collagen depends principally on two receptors, alpha (2)beta(1) integrin (GPIa-IIa) and GPVI. During this activation, the nonrec eptor protein tyrosine kinase pp72(syk) is rapidly phosphorylated, but the precise contribution of alpha(2)beta(1) integrin and GPVI to signaling for this phosphorylation is not clear. We have recently found that proteolysis of platelet alpha(2)beta(1) integrin by the snake venom metalloproteinase, jararhagin, results in inhibition of collagen-induced platelet aggregation and pp72(syk) phosphorylation, In order to verify whether the treatment of platelets with jararhagin had any effect on GPVI signaling, in this study w e stimulated platelets treated with either jararhagin or anti-alpha(2)beta( 1) antibody with two GPVI agonists, an antibody to GPVI and convulxin. Plat elet shape change and phosphorylation of pp72(syk) by both GPVI agonists wa s preserved, as was the structure and function of GPVI shown by I-125-label ed convulxin binding to immunoprecipitated GPVI from jararhagin-treated pla telets. In contrast, defective platelet aggregation in response to GPVI ago nists occurred in both jararhagin-treated and alpha(2)beta(1)-blocked plate lets. This apparent cosignaling role of alpha(2)beta(1) integrin for platel et aggregation suggests the possibility of a topographical association of t his integrin with GPVI, We found that both platelet alpha(2)beta(1) integri n and GPVI coimmunoprecipitated with alpha(2)beta(1) integrin, Since platel et aggregation requires activation of alpha(IIb)beta(3) integrin, defective aggregation in the absence of alpha(2)beta(1) suggests that this receptor may provide a signaling link between GPVI and alpha(IIb)beta(3). Our study therefore demonstrates that platelet signaling leading to pp72(syk) phospho rylation initiated with GPVI engagement by either convulxin or GPVI antibod y does not depend on alpha(2)beta(1) integrin. However, alpha(IIb)beta(3), integrin may, in this model, require functional alpha(2)beta(1) integrin fo r its activation. (C) 2000 Academic Press.