Evidence of a role for SHP-1 in platelet activation by the collagen receptor glycoprotein VI

The Src homology (SH)2 domain-containing protein-tyrosine phosphatase SHP-1 is tyrosine phosphorylated in platelets in response to the glycoprotein VI (GPVI)-selective agonist collagen-related peptide (CRP), collagen, and thr ombin. Two major unidentified tyrosine-phosphorylated bands of 28 and 32 kD a and a minor band of 130 kDa coprecipitate with SHP-1 in response to all t hree agonists. Additionally, tyrosine-phosphorylated proteins of 50-55 and 70 kDa specifically associate with SHP-1 following stimulation by CRP and c ollagen. The tyrosine kinases Lyn, which exists as a 53 and 56-kDa doublet, and Syk were identified as major components of these bands, respectively. Kinase assays on SHP-1 immunoprecipitates performed in the presence of the Src family kinase inhibitor PP1 confirmed the presence of a Src kinase in C RP- but not thrombin-stimulated cells. Lyn, Syk, and SLP-76, along with tyr osine-phosphorylated 28-, 32-, and 130-kDa proteins, bound selectively to a glutathione S-transferase protein encoding the SH2 domains of SHP-1, sugge sting that this is the major site of interaction. Platelets isolated from m otheaten viable mice (mev/mev) revealed the presence of a heavily tyrosine- phosphorylated 26-kDa protein that was not found in wild-type platelets. CR P-stimulated mev/mev platelets manifested hypophosphorylation of Syk and Ly n and reduced P-selectin expression relative to controls. These observation s provide evidence of a functional role for SHP-1 in platelet activation by GPVI.