Grb2 promotes integrin-induced focal adhesion kinase (FAK) autophosphorylation and directs the phosphorylation of protein tyrosine phosphatase α by the Src-FAK kinase complex

Mol Cell Biol. 2014 Feb;34(3):348-61. doi: 10.1128/MCB.00825-13. Epub 2013 Nov 18.

Abstract

The integrin-activated Src-focal adhesion kinase (FAK) kinase complex phosphorylates PTPα at Tyr789, initiating PTPα-mediated signaling that promotes cell migration. Recruitment of the BCAR3-Cas complex by PTPα-phospho-Tyr789 at focal adhesions is one mechanism of PTPα signaling. The adaptor protein Grb2 is also recruited by PTPα-phospho-Tyr789, although the role of the PTPα-Grb2 complex in integrin signaling is unknown. We show that silencing Grb2 expression in fibroblasts abolishes PTPα-Tyr789 phosphorylation and that this is due to two unexpected actions of Grb2. First, Grb2 promotes integrin-induced autophosphorylation of FAK-Tyr397. This is impaired in Grb2-depleted cells and prohibits FAK activation and formation of the Src-FAK complex. Grb2-depleted cells contain less paxillin, and paxillin overexpression rescues FAK-Tyr397 phosphorylation, suggesting that the FAK-activating action of Grb2 involves paxillin. A second distinct role for Grb2 in PTPα-Tyr789 phosphorylation involves Grb2-mediated coupling of Src-FAK and PTPα. This requires two phosphosites, FAK-Tyr925 and PTPα-Tyr789, for Grb2-Src homology 2 (SH2) binding. We propose that a Grb2 dimer links FAK and PTPα, and this positions active Src-FAK in proximity with other, perhaps integrin-clustered, molecules of PTPα to enable maximal PTPα-Tyr789 phosphorylation. These findings identify Grb2 as a new FAK activator and reveal its essential role in coordinating PTPα tyrosine phosphorylation to enable downstream integrin signaling and migration.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cells, Cultured
  • Embryo, Mammalian / cytology
  • Fibroblasts / cytology
  • Fibroblasts / metabolism*
  • Focal Adhesion Kinase 1 / genetics
  • Focal Adhesion Kinase 1 / metabolism*
  • GRB2 Adaptor Protein / genetics
  • GRB2 Adaptor Protein / metabolism*
  • Immunoblotting
  • Integrins / metabolism
  • Mice
  • Mice, Knockout
  • Paxillin / genetics
  • Paxillin / metabolism
  • Phosphorylation
  • Protein Binding
  • RNA Interference
  • Receptor-Like Protein Tyrosine Phosphatases, Class 4 / genetics
  • Receptor-Like Protein Tyrosine Phosphatases, Class 4 / metabolism*
  • Signal Transduction
  • Tyrosine / genetics
  • Tyrosine / metabolism
  • src Homology Domains*

Substances

  • GRB2 Adaptor Protein
  • Grb2 protein, mouse
  • Integrins
  • Paxillin
  • Tyrosine
  • Focal Adhesion Kinase 1
  • Ptk2 protein, mouse
  • Ptpra protein, mouse
  • Receptor-Like Protein Tyrosine Phosphatases, Class 4