Pyk2 and Src-family protein-tyrosine kinases compensate for the loss of FAK in fibronectin-stimulated signaling events but Pyk2 does not fully function to enhance FAK- cell migration

EMBO J. 1998 Oct 15;17(20):5933-47. doi: 10.1093/emboj/17.20.5933.

Abstract

The focal adhesion kinase (FAK) protein-tyrosine kinase (PTK) links transmembrane integrin receptors to intracellular signaling pathways. We show that expression of the FAK-related PTK, Pyk2, is elevated in fibroblasts isolated from murine fak-/- embryos (FAK-) compared with cells from fak+/+ embryos (FAK+). Pyk2 was localized to perinuclear regions in both FAK+ and FAK- cells. Pyk2 tyrosine phosphorylation was enhanced by fibronectin (FN) stimulation of FAK- but not FAK+ cells. Increased Pyk2 tyrosine phosphorylation paralleled the time-course of Grb2 binding to Shc and activation of ERK2 in FAK- cells. Pyk2 in vitro autophosphorylation activity was not enhanced by FN plating of FAK- cells. However, Pyk2 associated with active Src-family PTKs after FN but not poly-L-lysine replating of the FAK- cells. Overexpression of both wild-type (WT) and kinase-inactive (Ala457), but not the autophosphorylation site mutant (Phe402) Pyk2, enhanced endogenous FN-stimulated c-Src in vitro kinase activity in FAK- cells, but only WT Pyk2 overexpression enhanced FN-stimulated activation of co-transfected ERK2. Interestingly, Pyk2 overexpression only weakly augmented FAK- cell migration to FN whereas transient FAK expression promoted FAK- cell migration to FN efficiently compared with FAK+ cells. Significantly, repression of endogenous Src-family PTK activity by p50(csk) overexpression inhibited FN-stimulated cell spreading, Pyk2 tyrosine phosphorylation, Grb2 binding to Shc, and ERK2 activation in the FAK- but not in FAK+ cells. These studies show that Pyk2 and Src-family PTKs combine to promote FN-stimulated signaling events to ERK2 in the absence of FAK, but that these signaling events are not sufficient to overcome the FAK- cell migration defects.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Calcium-Calmodulin-Dependent Protein Kinases / physiology
  • Cell Adhesion Molecules / analysis*
  • Cell Adhesion Molecules / physiology*
  • Cell Movement / physiology*
  • Cells, Cultured
  • Fibroblasts
  • Fibronectins / metabolism*
  • Focal Adhesion Kinase 1
  • Focal Adhesion Kinase 2
  • Focal Adhesion Protein-Tyrosine Kinases
  • Humans
  • Mice
  • Mitogen-Activated Protein Kinase 1
  • Molecular Sequence Data
  • Protein-Tyrosine Kinases / analysis*
  • Protein-Tyrosine Kinases / biosynthesis
  • Protein-Tyrosine Kinases / deficiency*
  • Protein-Tyrosine Kinases / physiology*
  • Rabbits
  • Signal Transduction / physiology*
  • Vinculin / analysis
  • src-Family Kinases / physiology*

Substances

  • Cell Adhesion Molecules
  • Fibronectins
  • Vinculin
  • Protein-Tyrosine Kinases
  • Focal Adhesion Kinase 1
  • Focal Adhesion Kinase 2
  • Focal Adhesion Protein-Tyrosine Kinases
  • PTK2 protein, human
  • Ptk2 protein, mouse
  • Ptk2b protein, mouse
  • src-Family Kinases
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Mitogen-Activated Protein Kinase 1