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Paxillin null embryonic stem cells are impaired in cell spreading and tyrosine phosphorylation of focal adhesion kinase

Abstract

Paxillin is a focal-adhesion associated protein implicated in the regulation of integrin signaling and organization of the actin cytoskeleton. Paxillin associates with numerous signaling molecules including adaptor molecules (p130Cas, CRK), kinases (FAK, Pyk2, PAK and SRC), tyrosine phosphatases (PTP–PEST), ARF–GAP proteins (p95pkl, PAG3) and papillomavirus E6 oncoproteins. Although paxillin is tyrosine phosphorylated in cellular processes such as cell attachment and spreading, little direct evidence is available about paxillin's role in these events. Targeted gene disruption was used to generate paxillin null mouse embryonic stem (ES) cells and paxillin null differentiated cells. Paxillin null ES cells exhibit delayed spreading on integrin binding substrates fibronectin and laminin, and there is reduced tyrosine phosphorylation of Focal Adhesion Kinase (FAK). Both of these phenotypes are recovered in paxillin knockout cells upon exogenous re-expression of paxillin. The individual LD motifs of paxillin that are binding sites for FAK, vinculin and ARF–GAP proteins, as well as tyrosine residues that when phosphorylated create binding sites for CRK family members, are dispensable for FAK phosphorylation and early cell spreading. These results demonstrate that paxillin contributes to attachment-dependent tyrosine phosphorylation of FAK and early cell spreading in ES cells.

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Acknowledgements

The authors wish to thank Dr Chris Turner for providing the paxillin wild type cDNA and the 1–331 mutant of paxillin, and Drs Ron Conlon and David LePage for the generous gifts of the ES cells, targeting vectors, and helpful advise. The authors also thank Drs Lloyd Culp and Susann Brady-Kalnay for critical reading of this manuscript. This work was supported by NIH grant CA69292 to S Vande Pol.

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Correspondence to Scott Vande Pol.

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Wade, R., Bohl, J. & Vande Pol, S. Paxillin null embryonic stem cells are impaired in cell spreading and tyrosine phosphorylation of focal adhesion kinase. Oncogene 21, 96–107 (2002). https://doi.org/10.1038/sj.onc.1205013

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  • DOI: https://doi.org/10.1038/sj.onc.1205013

Keywords

  • cytoskeleton
  • paxillin
  • FAK
  • integrin
  • kinases
  • actin

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