Chu P.-Y.Huang L.-Y.Hsu C.-H.Liang C.-C.Guan J.-L.Hung T.-H.TING-HSUAN HUNGHsu, Chun-HuaChun-HuaHsuShen, Tang-LongTang-LongShen2020-01-022020-01-02200900219258https://scholars.lib.ntu.edu.tw/handle/123456789/444377We have previously reported that growth factor receptor-bound protein-7 (Grb7), an Src-homology 2 (SH2)-containing adaptor protein, enables interaction with focal adhesion kinase (FAK) to regulate cell migration in response to integrin activation. To further elucidate the signaling events mediated by FAK-Grb7 complexes in promoting cell migration and other cellular functions, we firstly examined the phosphorylated tyrosine site(s) of Grb7 by FAK using an in vivo mutagenesis. We found that FAK was capable of phosphorylating at least 2 of 12 tyrosine residues within Grb7, Tyr-188 and Tyr-338. Moreover, mutations converting the identified Tyr to Phe inhibited integrin-dependent cell migration as well as impaired cell proliferation but not survival compared with the wild-type control. Interestingly, the above inhibitory effects caused by the tyrosine phosphorylation-deficient mutants are probably attributed to their down-regulation of phospho-Tyr-397 of FAK, thereby implying a mechanism by competing with wild-type Grb7 for binding to FAK. Consequently, these tyrosine phosphorylation-deficient mutants evidently altered the phospho-Tyr-118 of paxillin and phosphorylation of ERK1/2 but less on phospho-Ser-473 of AKT, implying their involvement in the FAK-Grb7-mediated cellular functions. Additionally, we also illustrated that the formation of FAK-Grb7 complexes and Grb7 phosphorylation by FAK in an integrin-dependent manner were essential for cell migration, proliferation and anchorage-independent growth in A431 epidermal carcinoma cells, indicating the importance of FAK-Grb7 complexes in tumorigenesis. Our data provide a better understanding on the signal transduction event for FAK-Grb7-mediated cellular functions as well as to shed light on a potential therapeutic in cancers. ? 2009 by The American Society for Biochemistry and Molecular Biology, Inc.[SDGs]SDG3Adaptor proteins; Bound proteins; Carcinoma cells; Cell migration; Cellular function; Down-regulation; Focal adhesion kinase; Growth factor; In-vivo; Inhibitory effect; Integrin; Integrin activation; Paxillin; Phosphorylated tyrosine; Signaling events; Src-homology; Tumorigenesis; Tyrosine phosphorylation; Tyrosine residues; Wild types; Wild-type controls; Adhesion; Amino acids; Bioactivity; Cell membranes; Cell proliferation; Peptides; Signal transduction; Phosphorylation; focal adhesion kinase; growth factor receptor bound protein 7; integrin; mitogen activated protein kinase 1; mutant protein; paxillin; phenylalanine; protein kinase B; serine; tyrosine; animal cell; article; carcinogenesis; carcinoma cell; cell anchorage; cell function; cell growth; cell migration; cell proliferation; cell survival; controlled study; down regulation; human; human cell; molecular mechanics; mutagenesis; nonhuman; priority journal; protein binding; protein phosphorylation; protein protein interaction; regulatory mechanism; signal transduction; wild type; Animals; Carcinoma; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; CHO Cells; Cricetinae; Cricetulus; Extracellular Signal-Regulated MAP Kinases; Focal Adhesion Protein-Tyrosine Kinases; Gene Expression Regulation; GRB7 Adaptor Protein; Humans; Integrins; Mice; NIH 3T3 Cells; Paxillin; Phosphorylation; Point Mutation; Proto-Oncogene Proteins c-akt; Tyrosinejournal article10.1074/jbc.M109.0182592-s2.0-67749099777https://www2.scopus.com/inward/record.uri?eid=2-s2.0-67749099777&doi=10.1074%2fjbc.M109.018259&partnerID=40&md5=64df29c4fd79f821687f92acd57c8939