Integrin-dependent PLC-gamma1 phosphorylation mediates fibronectin-dependent adhesion.
Although integrin engagement initiates signaling events such as focal-adhesion kinase (FAK) and Src kinase activation, the role of phosphoinositide turnover in cell adhesion is less clear. To assess PLC-gamma1 function in this process, Plcg1(-/-) fibroblasts (Null) were compared with the same fibroblasts in which PLC-gamma1 was re-expressed (Null+). Following plating on fibronectin, Null cells displayed a significantly impaired rate of adhesion compared with Null+ cells. This defect was detected at low concentrations of fibronectin; at high fibronectin concentrations, the Null and Null+ cells displayed equivalent adhesion characteristics. The differences were not due to PLC-gamma1-dependent changes in integrin subunit expression, nor was integrin receptor clustering impaired with the absence of PLC-gamma1. Experiments with site-specific antibodies and PLC-gamma1 mutants showed that fibronectin selectively increased phosphorylation of Tyr783 and that mutagenesis of this residue, but not Tyr771 or Tyr1253, abrogated fibronectin-dependent adhesion. The SH2 domains of PLC-gamma1 were also required for maximal adhesion on fibronectin. Adhesion to fibronectin induced PLC-gamma1 tyrosine phosphorylation that was inhibited by a Src-kinase inhibitor, but not an epidermal-growth-factor-receptor kinase inhibitor. Moreover, in cells null for Src family members, but not in cells null for FAK family members, integrin-dependent PLC-gamma1 tyrosine phosphorylation was greatly reduced. Finally, the data demonstrated that PLC-gamma1 co-immunoprecipitated with Src following fibronectin-induced integrin activation, and this association did not depend on FAK expression.
Tvorogov
Denis
D
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
Wang
Xue-Jie
XJ
Zent
Roy
R
Carpenter
Graham
G
D000818:Animals; D002448:Cell Adhesion / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002465:Cell Movement / Q000235:genetics / Q000502:physiology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019012:Integrin beta1 / Q000378:metabolism; D016023:Integrins / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D051966:Phospholipase C gamma; D010766:Phosphorylation; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D010738:Type C Phospholipases / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism
D005353:Fibronectins; D019012:Integrin beta1; D016023:Integrins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D019061:src-Family Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma