A Collagen-Like Peptide Stimulates Tyrosine Phosphorylation of syk and Phospholipase Cγ2 in Platelets Independent of the Integrin α2β1

Abstract

Activation of platelets by collagen is mediated through a tyrosine kinase-dependent pathway that is associated with phosphorylation of the Fc receptor γ chain, the tyrosine kinase syk, and phospholipase Cγ2 (PLCγ2). We recently described a collagen-related triple-helical peptide (CRP) with the sequence GCP*(GPP*)GCP*G (single letter amino acid code: P* = hydroxyproline; Morton et al, Biochem J 306:337, 1995). The cross-linked peptide is a potent stimulus of platelet activation but, unlike collagen, does not support α₂β₁-mediated, Mg²⁺-dependent adhesion, suggesting that its action is independent of the integrin α₂β₁ . This finding suggests the existence of a platelet receptor other than α₂β₁ that underlies activation. In the present study, we show that CRP stimulates tyrosine phosphorylation of the same pattern of proteins in platelets as collagen, including syk and PLCγ2. Protein tyrosine phosphorylation induced by CRP is not altered in the absence of Mg²⁺ or the presence of monoclonal antibodies (MoAbs) to the integrin α₂β₁ (MoAb 6F1 and MoAb 13), conditions that prevent the interaction of collagen with the integrin. In contrast, phosphorylation of syk and PLCγ2 by collagen is partially reduced by MoAb 6F1 and MoAb 13 or by removal of Mg²⁺. This may reflect a direct role of α₂β₁ in collagen-induced signaling events or an indirect role in which the integrin facilitates the binding of collagen to its signaling receptor. The results show an α₂β₁-independent pathway of platelet activation by CRP that involves phosphorylation of syk and PLCγ2. This pathway appears to contribute to platelet activation by collagen.