The cytoplasmic tyrosines of integrin subunit β1 are involved in focal adhesion kinase activation

We have previously shown that mutation of the two tyrosines in the cytoplasmic domain of integrin subunit beta 1 (Y783 and Y795) to phenylalanines markedly reduces the capability of beta 1A integrins to mediate directed cell migration. In this study, beta 1-dependent cell spreading was found to be delayed in GD25 cells expressing beta 1A(Y783/795F) compared to that in wild-type GD25-beta 1A. Focal adhesion kinase (FAK) tyrosine phosphorylation and activation were severely impaired in response to beta 1-dependent adhesion in GD25-beta 1A(Y783/795F) cells compared to that in wild-type GD25-beta 1A or mutants in which only a single tyrosine was altered (beta 1A(Y783F) or beta 1A(Y795F)). Phosphorylation site-specific antibodies selective for FAK phosphotyrosine 397 indicated that the defect in FAK phosphorylation via beta 1A(Y783/795F) lies at the level of the initial autophosphorylation step. Indeed, beta 1A-dependent tyrosine phosphorylation of tensin and paxillin was lost in the beta 1A(Y783/795F) cells, consistent with the impairment in FAK activation. in contrast, p130(CAS) overall tyrosine phosphorylation was unaffected by the beta 1 mutations. Despite the defect in beta 1-mediated FAK activation, FAK was still localized to focal adhesions, Taken together, the phenotype of the GD25-beta 1A(Y783/795F) cells resembles, but is distinct from, the phenotype observed in FAK-null cells. These observations argue that tyrosines 783 and 795 within the cytoplasmic tail of integrin subunit beta 1A are critical mediators of FAK activation and cell spreading in GD25 cells.