Integrin α4β1 promotes focal adhesion kinase-independent cell motility via α4 cytoplasmic domain-specific activation of c-Src

The fibronectin binding integrins alpha 5 beta 1 and alpha 4 beta 1 generate signals pivotal for cell migration through distinct yet undefined mechanisms. For alpha 5 beta 1, beta 1-mediated activation of focal adhesion kinase (FAK) promotes c-Src recruitment to FAK and the formation of a FAK-Src signaling complex. Herein, we show that FAK expression is essential for alpha 5 beta 1-stimulated cell motility and that exogenous expression of human alpha 4 in FAK-null fibroblasts forms a functional alpha 4 beta 1 receptor that promotes robust cell motility equal to the alpha 5 beta 1 stimulation of wild-type and FAK-reconstituted fibroblasts. alpha 4 beta 1-stimulated FAK-null cell spreading and motility were dependent on the integrity of the alpha 4 cytoplasmic domain, independent of direct paxillin binding to alpha 4, and were not affected by PRNK expression, a dominant-negative inhibitor of Pyk2. alpha 4 cytoplasmic domain-initiated signaling led to a similar to 4-fold activation of c-Src which did not require paxillin binding to alpha 4. Notably, alpha 4-stimulated cell motility was inhibited by catalytically inactive receptor protein-tyrosine phosphatase a overexpression and blocked by the p50Csk phosphorylation of c-Src at Tyr-529. (alpha 4 beta 1-stimulated cell motility of triple-null Src(-/-), c-Yes(-/-), and Fyn(-/-) fibroblasts was dependent on c-Src reexpression that resulted in p130Cas tyrosine phosphorylation and Rac GTPase loading. As p130as phosphorylation and Rac activation are common downstream targets for alpha 5 beta 1-stimulated FAK activation, our results support the existence of a novel alpha 4 cytoplasmic domain connection leading to c-Src activation which functions as a FAK-independent linkage to a common motility-promoting signaling pathway.