Induced focal adhesion kinase (FAK) expression in FAK-null cells enhances cell spreading and migration requiring both auto- and activation loop phosphorylation sites and inhibits adhesion-dependent tyrosine phosphorylation of Pyk2.
Focal adhesion kinase (FAK) is a nonreceptor protein tyrosine kinase involved in integrin-mediated control of cell behavior. Following cell adhesion to components of the extracellular matrix, FAK becomes phosphorylated at multiple sites, including tyrosines 397, 576, and 577. Tyr-397 is an autophosphorylation site that promotes interaction with c-Src or Fyn. Tyr-576 and Tyr-577 lie in the putative activation loop of the kinase domain, and FAK catalytic activity may be elevated through phosphorylation of these residues by associated Src family kinase. Recent studies have implicated FAK as a positive regulator of cell spreading and migration. To further study the mechanism of adhesion-induced FAK activation and the possible role and signaling requirements for FAK in cell spreading and migration, we utilized the tetracycline repression system to achieve inducible expression of either wild-type FAK or phosphorylation site mutants in fibroblasts derived from FAK-null mouse embryos. Using these Tet-FAK cells, we demonstrated that both the FAK autophosphorylation and activation loop sites are critical for maximum adhesion-induced FAK activation and FAK-enhanced cell spreading and migration responses. Negative effects on cell spreading and migration, as well as decreased phosphorylation of the substrate p130(Cas), were observed upon induced expression of the FAK autophosphorylation site mutant. These negative effects appear to result from an inhibition of integrin-mediated signaling by the FAK-related kinase Pyk2/CAKbeta/RAFTK/CadTK.
Owen
J D
JD
Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Ruest
P J
PJ
Fry
D W
DW
Hanks
S K
SK
D016475:3T3 Cells; D000818:Animals; D001665:Binding Sites; D002384:Catalysis; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D002465:Cell Movement / Q000502:physiology*; D050719:Crk-Associated Substrate Protein; D005353:Fibronectins / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051418:Focal Adhesion Kinase 2; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015870:Gene Expression; D051379:Mice; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D011506:Proteins; D011817:Rabbits; D050717:Retinoblastoma-Like Protein p130; D013752:Tetracycline / Q000494:pharmacology; D013997:Time Factors; D014443:Tyrosine / Q000378:metabolism*; D015854:Up-Regulation
C493842:Bcar1 protein, mouse; D015815:Cell Adhesion Molecules; D050719:Crk-Associated Substrate Protein; D005353:Fibronectins; D010750:Phosphoproteins; D011506:Proteins; D050717:Retinoblastoma-Like Protein p130; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051418:Focal Adhesion Kinase 2; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; C496053:Ptk2b protein, mouse; D013752:Tetracycline