Integrin α4β1 Promotes Focal Adhesion Kinase-Independent Cell Motility via α4 Cytoplasmic Domain-Specific Activation of c-Src

Abstract

The fibronectin binding integrins α5β1 and α4β1 generate signals pivotal for cell migration through distinct yet undefined mechanisms. For α5β1, β1-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 α5β1-stimulated cell motility and that exogenous expression of human α4 in FAK-null fibroblasts forms a functional α4β1 receptor that promotes robust cell motility equal to the α5β1 stimulation of wild-type and FAK-reconstituted fibroblasts. α4β1-stimulated FAK-null cell spreading and motility were dependent on the integrity of the α4 cytoplasmic domain, independent of direct paxillin binding to α4, and were not affected by PRNK expression, a dominant-negative inhibitor of Pyk2. α4 cytoplasmic domain-initiated signaling led to a ∼4-fold activation of c-Src which did not require paxillin binding to α4. Notably, α4-stimulated cell motility was inhibited by catalytically inactive receptor protein-tyrosine phosphatase α overexpression and blocked by the p50Csk phosphorylation of c-Src at Tyr-529. α4β1-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 p130Cas phosphorylation and Rac activation are common downstream targets for α5β1-stimulated FAK activation, our results support the existence of a novel α4 cytoplasmic domain connection leading to c-Src activation which functions as a FAK-independent linkage to a common motility-promoting signaling pathway.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/.

ACKNOWLEDGMENTS

We thank Martin Hemler for providing α4 integrin cDNAs and Jaewon Han for polyclonal antibody to human α4 integrin, and we greatly appreciate the administrative assistance provided by Theresa Villalpando.

S. Mitra was supported by a fellowship (12FT-0122) from the California Tobacco-Related Disease Research Program. D. N. Streblow was supported by an American Heart Association Scientist Development Award. This work was supported by grants from the NIH to Dusko Ilic (CA087652), Mark Ginsberg (AR027214), and David Schlaepfer (CA75240, CA87038, CA102310). David Schlaepfer is an Established Investigator of the American Heart Association (0540115N). This is manuscript number 17162-IMM from The Scripps Research Institute.