Nckbeta 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 Nckalpha (Nck) and Nckbeta (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 Nckbeta in platelet-derived growth factor (PDGF)-induced actin polymerization in NIH 3T3 cells. Overexpression of Nckbeta but not Nckalpha blocks PDGF-stimulated membrane ruffling and formation of lamellipoda. Mutation in either the SH2 or the middle SH3 domain of Nckbeta abolishes its interfering effect. Nckbeta binds at Tyr-1009 in human PDGF receptor beta (PDGFR-beta) which is different from Nckalpha's binding site, Tyr-751, and does not compete with phosphatidylinositol-3 kinase for binding to PDGFR. Microinjection of an anti-Nckbeta but not an anti-Nckalpha antibody inhibits PDGF-stimulated actin polymerization. Constitutively membrane-bound Nckbeta but not Nckalpha blocks Rac1-L62-induced membrane ruffling and formation of lamellipodia, suggesting that Nckbeta acts in parallel to or downstream of Rac1. This is the first report of Nckbeta's role in receptor tyrosine kinase signaling to the actin cytoskeleton.
Chen
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Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois 60637, USA.
She
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Kim
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Woodley
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DT
Li
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D016475:3T3 Cells; D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000077214:Becaplermin; D015152:Blotting, Northern; D015153:Blotting, Western; D002458:Cell Fractionation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004285:Dogs; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007668:Kidney / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D016297:Mutagenesis, Site-Directed; D015513:Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D020033:Protein Isoforms; D017368:Protein Prenylation; D020574:Proto-Oncogene Proteins c-sis; D011554:Pseudopodia / Q000378:metabolism; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D018909:src Homology Domains
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; C078332:Nck protein; D015513:Oncogene Proteins; D010982:Platelet-Derived Growth Factor; D020033:Protein Isoforms; D020574:Proto-Oncogene Proteins c-sis; D000077214:Becaplermin; D014443:Tyrosine; D019869:Phosphatidylinositol 3-Kinases; D020797:Receptor, Platelet-Derived Growth Factor beta; D020830:rac1 GTP-Binding Protein