Nckβ adapter regulates actin polymerization in NIH 3T3 fibroblasts in response to platelet-derived growth factor bb

The SH3-SH3-SH3-SH2 adapter Nck represents a two-gene family that includes Nck alpha (Nck) and Nck beta (Grb4/Nck2), and it links receptor tyrosine kinases to intracellular signaling networks. The function of these mammalian Nck genes has not been established. We report here a specific role for Nck beta in platelet-derived growth factor (PDGF)-induced actin polymerization in NIH 3T3 cells. Overexpression of Nck beta but not Nck alpha blocks PDGF-stimulated membrane ruffling and formation of lamellipoda. Mutation in either the SH2 or the middle SH3 domain of Nck beta abolishes its interfering effect. Nck beta binds at Tyr-1009 in human PDGF receptor beta (PDGFR-beta) which is different from Nck alpha's binding site, Tyr-751, and does not compete with phosphatidylinositol-3 kinase for binding to PDGFR Microinjection of an anti-Nck beta but not an anti-Nck alpha antibody inhibits PDGF-stimulated actin polymerization. Constitutively membrane-bound Nck beta but not Nck alpha blocks Rac1-L62-induced membrane ruffling and formation of lamellipodia, suggesting that Nck beta acts in parallel to or downstream of Rac1. This is the first report of Nck beta's role in receptor tyrosine kinase signaling to the actin cytoskeleton.