Interaction of linker for activation of T cells with multiple adapter proteins in platelets activated by the glycoprotein VI-selective ligand, convulxin

The snake venom toxin convulxin activates platelets through the collagen re ceptor glycoprotein VI (GPVI)/Fc receptor gamma -chain (FcR gamma -chain) c omplex leading to tyrosine phosphorylation and activation of the tyrosine S yk and phospholipase C gamma2 (PLC gamma2), In the present study, we demons trate that convulxin is a considerably more powerful agonist than collagen or the GPVI-selective collagen-related peptide (CRP), Confirmation that the response to convulxin is mediated solely via Syk was provided by studies o n Syk-deficient platelets. The increase in phosphorylation of the FcR gamma -chain is associated with marked increases in tyrosine phosphorylation of downstream proteins including Syk, linker for activation of T cells (LAT), SLP-76, and PLC gamma2, The transmembrane adapter LAT coprecipitates with S LP-76 and PLC gamma2, as well as with a number of other adapter proteins, s ome of which have not been previously described in platelets, including Cbl , Grb2, Gads, and SKAP-HOM. Cads is constitutively associated with SLP-76 a nd is probably the protein bridging its association with LAT, There was no detectable association between Grb2 and SLP-76 in control or stimulated cel ls, suggesting that the interaction of LAT with Grb2 is present in a separa te complex to that of LAT Gads SLP-76. These results show that the trimeric convulxin stimulates a much greater phosphorylation of the FcR gamma -chai n and subsequent downstream responses relative to CRP and collagen, presuma bly because of its ability to cause a greater degree of cross-linking of GP VI, The adapter LAT appears to play a critical role in recruiting a number of other adapter proteins to the surface membrane in response to activation of GPVI, presumably at sites of glycolipid-enriched microdomains, enabling an organized signaling cascade that leads to platelet activation.