Inhibition of cell spreading by expression of the C-terminal domain of focal adhesion kinase (FAK) is rescued by coexpression of Src or catalytically inactive FAK: a role for paxillin tyrosine phosphorylation.
pp125FAK is a tyrosine kinase that appears to regulate the assembly of focal adhesions and thereby promotes cell spreading on the extracellular matrix. In some cells, the C terminus of pp125FAK is expressed as a separate protein, pp41/43FRNK. We have previously shown that overexpression of pp41/43FRNK inhibits tyrosine phosphorylation of pp125FAK and paxillin and, in addition, delays cell spreading and focal adhesion assembly. Thus, pp41/43FRNK functions as a negative inhibitor of adhesion signaling and provides a tool to dissect the mechanism by which pp125FAK promotes cell spreading. We report here that the inhibitory effects of pp41/43FRNK expression can be rescued by the co-overexpression of wild-type pp125FAK and partially rescued by catalytically inactive variants of pp125FAK. However, coexpression of an autophosphorylation site mutant of pp125FAK, which fails to bind the SH2 domain of pp60c-Src, or a mutant that fails to bind paxillin did not promote cell spreading. In contrast, expression of pp41/43FRNK and pp60c-Src reconstituted cell spreading and tyrosine phosphorylation of paxillin but did so without inducing tyrosine phosphorylation of pp125FAK. These data provide additional support for a model whereby pp125FAK acts as a "switchable adaptor" that recruits pp60c-Src to phosphorylate paxillin, promoting cell spreading. In addition, these data point to tyrosine phosphorylation of paxillin as being a critical step in focal adhesion assembly.
Richardson
A
A
Department of Microbiology, Health Sciences Center, University of Virginia, Charlottesville 22908, USA.
Malik
R K
RK
Hildebrand
J D
JD
Parsons
J T
JT
D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion / Q000502:physiology*; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015870:Gene Expression; D008954:Models, Biological; D009154:Mutation; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism
D015815:Cell Adhesion Molecules; D003598:Cytoskeletal Proteins; D051419:Paxillin; D010750:Phosphoproteins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D016392:Proto-Oncogene Proteins pp60(c-src)