Grb2 negatively regulates epidermal growth factor-induced phospholipase C-γ1 activity through the direct interaction with tyrosine-phosphorylated phospholipase C-γ1

Phospholipase C-gamma 1 (PLC-gamma 1). plays pivotal roles in cellular growth and proliferation. Upon the stimulation of growth factors and hormones, PLC-gamma 1 is rapidly phosphorylated at three known sites; Tyr(771), Tyr(783) and Tyr(1254) and its enzymatic activity is up-regulated. In this study, we demonstrate for the first time that Grb2, an adaptor protein, specifically interacts with tyrosine-phosphorylated,PLC-gamma 1 at Tyr(783). The association of Grb2 with PLC-gamma 1 was induced by the treatment with epidermal growth factor (EGF). Replacement of Tyr(783) with Phe completely blocked EGF-induced interaction of PLC-gamma 1 with Grb2, indicating that tyrosine phosphorylation of PLC-gamma 1 at Tyr783 is essential for the interaction with Grb2. Interestingly, the depletion of Grb2 from HEK-293 cells by RNA interference significantly enhanced increased EGF-induced PLC-gamma 1 enzymatic activity and mobilization of the intracellular Ca2+, while it did not affect EGF-induced tyrosine phosphorylation of PLC-gamma 1. Furthermore, overexpression of Grb2 inhibited PLC-gamma 1 enzymatic activity. Taken together, these results suggest Grb2, in addition to its key function in signaling through Ras, may have a negatively regulatory role on EGF-induced PLC-gamma 1 activation. (c) 2005 Elsevier Inc. All rights reserved.