CHARACTERIZATION OF GRB2-BINDING PROTEINS IN HUMAN PLATELETS ACTIVATED BY FC-GAMMA-RIIA CROSS-LINKING

Glutathione-S-transferase (GST)-Grb2 fusion proteins have been used go identify the potential role of Grb2-binding proteins in platelet acti vation by the platelet low-affinity IBG receptor, Fc gamma RIIA. Two t yrosine phosphoproteins of 38 and 63 kD bind to the SH2 domain of Grb2 following Fc gamma RIIA stimulation of platelets. Both are located in the particulate fraction Following platelet activation and are also a ble 40 bind to a GST-construct containing the SH2 and SH3 domains of p hospholipase C gamma 1. p38 also forms a complex with the tyrosine kin ase csk in stimulated cells and is a substrate for the kinase. The SH3 domains of Grb(2) form a stable complex with SOS1 and two proteins of 75 kD and 120 kD, which undergo tyrosine phosphorylation in Fc gamma RIIA-stimulated cells, The 75-kD protein is recognized by antibodies t o SLP-76, which has recently been isolated from T cells and sequenced. Tyrosine phosphorylation of p38 and p63 is also observed in platelets stimulated by the tyrosine kinase-linked receptor agonist collagen an d by the G protein-coupled receptor agonist thrombin, although phospho rylation of SLP-76 is only observed in collagen-stimulated platelets. p38 and p63 may provide a docking sire for Grb2, thereby linking Grb2 SHE-binding proteins SOS1, SLP-76, and p120 to downstream signalling e vents. (C) 1996 by The American Society of Hematology.