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We analyzed the interaction of convulxin (Cvx), a 72-kDa protein isolated from the venom of Crotalus durissus terrificus, with human platelets. Cvx is a potent platelet agonist that induces an increase in the intracel-

IntroductionPlatelet endothelial cell adhesion molecule-1 (PECAM-1, CD31) is a 130-kd membrane-spanning glycoprotein whose expression is restricted to several hematopoietic cell types including platelets, monocytes, neutrophils, certain T lymphocytes, and vascular endothelial cells. [1][2][3][4] The functions of PECAM-1 are diverse and include angiogenesis, 5 vasculogenesis, 6 integrin regulation, 7,8 transendothelial migration of leukocytes, 9-11 and T-and B-cell antigen receptor function, 12,13 though the role of this molecule in platelets is unclear. When PECAM-1 was cloned, it was assigned to the family of cell adhesion molecules on the basis of structural similarities. 4 PECAM-1 is involved in adhesion, though much attention has been directed recently to studying its ability to participate in signal transduction. The cytoplasmic tail of PECAM-1 contains a conserved motif called an immunoreceptor tyrosine-based inhibitory motif (ITIM), which underlies its signaling properties and is shared by a growing family of inhibitory receptors. These include immunoglobulin G (IgG) receptor Fc␥RIIB, killer inhibitory receptors, and signal regulatory proteins, but PECAM-1 is the only ITIM-bearing receptor reported to be expressed in platelets. 14,15 Therefore, it has been proposed that PECAM-1 be assigned to the immunoglobulin-ITIM family of receptors. 14 The ligand-binding properties of PECAM-1 are complex. It has the capacity for homophilic interactions 3,16 and heterophilic interactions with a number of molecules that include integrin ␣ v ␤ 3 and CD38. 17,18 PECAM-1 becomes phosphorylated on tyrosine residues in response to a variety of stimuli that include PECAM-1 cross-linking, 19 activation of the high-affinity receptor for IgE (Fc⑀RI), 20 shear stress, 21 and oxidative stress. 22 Furthermore, we have recently reported that platelet activation through the collagen receptor glycoprotein VI (GPVI) and through thrombin receptors results in PECAM-1 tyrosine phosphorylation and that this is not dependent on platelet aggregation and secretion, 23 though tyrosine phosphorylation is enhanced by aggregation. 24 The tyrosine residues that become phosphorylated in PECAM-1 have been mapped and fall within the ITIM. 25 Phosphorylated ITIMs recruit signaling molecules, such as the tyrosine phosphatases SHP-1 and SHP-2, which bind to the motif through Src-homology 2 domain interactions. Indeed, both SHP-1 and SHP-2 have been shown to associate with tyrosine-phosphorylated PECAM-1, [24][25][26] and PECAM-1 ITIM phosphopeptides activate these phosphatases in vitro. 19,25,26 Generally, these protein tyrosine phosphatases exhibit inhibitory effects by counteracting tyrosine kinase-dependent pathways, though SHP-2 has been shown to positively regulate growth factor receptor signaling. 27 Immunoreceptor tyrosine-based activatory motif (ITAM)-bearing receptors have a critical place in the regulation of platelet function. 28 Indeed, the collagen receptor GPVI-FcR ␥-chain complex signals through an ITAM on the cytoplasmic tail of the FcR ...

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