Distinct Contributions of Glycoprotein VI and α2β1 Integrin to the Induction of Platelet Protein Tyrosine Phosphorylation and Aggregation

Platelet activation by collagen depends principally on two receptors, α2β1 integrin (GPIa-IIa) and GPVI. During this activation, the nonreceptor protein tyrosine kinase pp72syk is rapidly phosphorylated, but the precise contribution of α2β1 integrin and GPVI to signaling for this phosphorylation is not clear. We have recently found that proteolysis of platelet α2β1 integrin by the snake venom metalloproteinase, jararhagin, results in inhibition of collagen-induced platelet aggregation and pp72syk phosphorylation. In order to verify whether the treatment of platelets with jararhagin had any effect on GPVI signaling, in this study we stimulated platelets treated with either jararhagin or anti-α2β1 antibody with two GPVI agonists, an antibody to GPVI and convulxin. Platelet shape change and phosphorylation of pp72syk by both GPVI agonists was preserved, as was the structure and function of GPVI shown by 125I-labeled convulxin binding to immunoprecipitated GPVI from jararhagin-treated platelets. In contrast, defective platelet aggregation in response to GPVI agonists occurred in both jararhagin-treated and α2β1-blocked platelets. This apparent cosignaling role of α2β1 integrin for platelet aggregation suggests the possibility of a topographical association of this integrin with GPVI. We found that both platelet α2β1 integrin and GPVI coimmunoprecipitated with αIIbβ3 integrin. Since platelet aggregation requires activation of αIIbβ3 integrin, defective aggregation in the absence of α2β1 suggests that this receptor may provide a signaling link between GPVI and αIIbβ3. Our study therefore demonstrates that platelet signaling leading to pp72syk phosphorylation initiated with GPVI engagement by either convulxin or GPVI antibody does not depend on α2β1 integrin. However, αIIbβ3 integrin may, in this model, require functional α2β1 integrin for its activation.