• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Dec 1997; 17(12): 6906–6914.
PMCID: PMC232547

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.

Abstract

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.

Full Text

The Full Text of this article is available as a PDF (973K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Akasaka T, van Leeuwen RL, Yoshinaga IG, Mihm MC, Jr, Byers HR. Focal adhesion kinase (p125FAK) expression correlates with motility of human melanoma cell lines. J Invest Dermatol. 1995 Jul;105(1):104–108. [PubMed]
  • Bachelot C, Rameh L, Parsons T, Cantley LC. Association of phosphatidylinositol 3-kinase, via the SH2 domains of p85, with focal adhesion kinase in polyoma middle t-transformed fibroblasts. Biochim Biophys Acta. 1996 Mar 27;1311(1):45–52. [PubMed]
  • Bergman M, Joukov V, Virtanen I, Alitalo K. Overexpressed Csk tyrosine kinase is localized in focal adhesions, causes reorganization of alpha v beta 5 integrin, and interferes with HeLa cell spreading. Mol Cell Biol. 1995 Feb;15(2):711–722. [PMC free article] [PubMed]
  • Bretscher MS. Getting membrane flow and the cytoskeleton to cooperate in moving cells. Cell. 1996 Nov 15;87(4):601–606. [PubMed]
  • Burridge K, Chrzanowska-Wodnicka M. Focal adhesions, contractility, and signaling. Annu Rev Cell Dev Biol. 1996;12:463–518. [PubMed]
  • Burridge K, Fath K, Kelly T, Nuckolls G, Turner C. Focal adhesions: transmembrane junctions between the extracellular matrix and the cytoskeleton. Annu Rev Cell Biol. 1988;4:487–525. [PubMed]
  • Calalb MB, Polte TR, Hanks SK. Tyrosine phosphorylation of focal adhesion kinase at sites in the catalytic domain regulates kinase activity: a role for Src family kinases. Mol Cell Biol. 1995 Feb;15(2):954–963. [PMC free article] [PubMed]
  • Cary LA, Chang JF, Guan JL. Stimulation of cell migration by overexpression of focal adhesion kinase and its association with Src and Fyn. J Cell Sci. 1996 Jul;109(Pt 7):1787–1794. [PubMed]
  • Chen HC, Appeddu PA, Isoda H, Guan JL. Phosphorylation of tyrosine 397 in focal adhesion kinase is required for binding phosphatidylinositol 3-kinase. J Biol Chem. 1996 Oct 18;271(42):26329–26334. [PubMed]
  • Chen HC, Appeddu PA, Parsons JT, Hildebrand JD, Schaller MD, Guan JL. Interaction of focal adhesion kinase with cytoskeletal protein talin. J Biol Chem. 1995 Jul 14;270(28):16995–16999. [PubMed]
  • Chen HC, Guan JL. Association of focal adhesion kinase with its potential substrate phosphatidylinositol 3-kinase. Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):10148–10152. [PMC free article] [PubMed]
  • Cobb BS, Schaller MD, Leu TH, Parsons JT. Stable association of pp60src and pp59fyn with the focal adhesion-associated protein tyrosine kinase, pp125FAK. Mol Cell Biol. 1994 Jan;14(1):147–155. [PMC free article] [PubMed]
  • Gilmore AP, Romer LH. Inhibition of focal adhesion kinase (FAK) signaling in focal adhesions decreases cell motility and proliferation. Mol Biol Cell. 1996 Aug;7(8):1209–1224. [PMC free article] [PubMed]
  • Guinebault C, Payrastre B, Racaud-Sultan C, Mazarguil H, Breton M, Mauco G, Plantavid M, Chap H. Integrin-dependent translocation of phosphoinositide 3-kinase to the cytoskeleton of thrombin-activated platelets involves specific interactions of p85 alpha with actin filaments and focal adhesion kinase. J Cell Biol. 1995 May;129(3):831–842. [PMC free article] [PubMed]
  • Hanks SK, Calalb MB, Harper MC, Patel SK. Focal adhesion protein-tyrosine kinase phosphorylated in response to cell attachment to fibronectin. Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8487–8491. [PMC free article] [PubMed]
  • Harte MT, Hildebrand JD, Burnham MR, Bouton AH, Parsons JT. p130Cas, a substrate associated with v-Src and v-Crk, localizes to focal adhesions and binds to focal adhesion kinase. J Biol Chem. 1996 Jun 7;271(23):13649–13655. [PubMed]
  • Hildebrand JD, Schaller MD, Parsons JT. Identification of sequences required for the efficient localization of the focal adhesion kinase, pp125FAK, to cellular focal adhesions. J Cell Biol. 1993 Nov;123(4):993–1005. [PMC free article] [PubMed]
  • Hildebrand JD, Schaller MD, Parsons JT. Paxillin, a tyrosine phosphorylated focal adhesion-associated protein binds to the carboxyl terminal domain of focal adhesion kinase. Mol Biol Cell. 1995 Jun;6(6):637–647. [PMC free article] [PubMed]
  • Hildebrand JD, Taylor JM, Parsons JT. An SH3 domain-containing GTPase-activating protein for Rho and Cdc42 associates with focal adhesion kinase. Mol Cell Biol. 1996 Jun;16(6):3169–3178. [PMC free article] [PubMed]
  • Huttenlocher A, Sandborg RR, Horwitz AF. Adhesion in cell migration. Curr Opin Cell Biol. 1995 Oct;7(5):697–706. [PubMed]
  • Hynes RO. Integrins: versatility, modulation, and signaling in cell adhesion. Cell. 1992 Apr 3;69(1):11–25. [PubMed]
  • Ilić D, Kanazawa S, Furuta Y, Yamamoto T, Aizawa S. Impairment of mobility in endodermal cells by FAK deficiency. Exp Cell Res. 1996 Feb 1;222(2):298–303. [PubMed]
  • Ilić D, Furuta Y, Kanazawa S, Takeda N, Sobue K, Nakatsuji N, Nomura S, Fujimoto J, Okada M, Yamamoto T. Reduced cell motility and enhanced focal adhesion contact formation in cells from FAK-deficient mice. Nature. 1995 Oct 12;377(6549):539–544. [PubMed]
  • Kanner SB, Reynolds AB, Vines RR, Parsons JT. Monoclonal antibodies to individual tyrosine-phosphorylated protein substrates of oncogene-encoded tyrosine kinases. Proc Natl Acad Sci U S A. 1990 May;87(9):3328–3332. [PMC free article] [PubMed]
  • Kaplan KB, Swedlow JR, Morgan DO, Varmus HE. c-Src enhances the spreading of src-/- fibroblasts on fibronectin by a kinase-independent mechanism. Genes Dev. 1995 Jun 15;9(12):1505–1517. [PubMed]
  • Lauffenburger DA, Horwitz AF. Cell migration: a physically integrated molecular process. Cell. 1996 Feb 9;84(3):359–369. [PubMed]
  • Lipfert L, Haimovich B, Schaller MD, Cobb BS, Parsons JT, Brugge JS. Integrin-dependent phosphorylation and activation of the protein tyrosine kinase pp125FAK in platelets. J Cell Biol. 1992 Nov;119(4):905–912. [PMC free article] [PubMed]
  • Mitchison TJ, Cramer LP. Actin-based cell motility and cell locomotion. Cell. 1996 Feb 9;84(3):371–379. [PubMed]
  • Miyamoto S, Akiyama SK, Yamada KM. Synergistic roles for receptor occupancy and aggregation in integrin transmembrane function. Science. 1995 Feb 10;267(5199):883–885. [PubMed]
  • Miyamoto S, Teramoto H, Coso OA, Gutkind JS, Burbelo PD, Akiyama SK, Yamada KM. Integrin function: molecular hierarchies of cytoskeletal and signaling molecules. J Cell Biol. 1995 Nov;131(3):791–805. [PMC free article] [PubMed]
  • Nobes CD, Hall A. Rho, rac, and cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia. Cell. 1995 Apr 7;81(1):53–62. [PubMed]
  • Owens LV, Xu L, Craven RJ, Dent GA, Weiner TM, Kornberg L, Liu ET, Cance WG. Overexpression of the focal adhesion kinase (p125FAK) in invasive human tumors. Cancer Res. 1995 Jul 1;55(13):2752–2755. [PubMed]
  • Palecek SP, Loftus JC, Ginsberg MH, Lauffenburger DA, Horwitz AF. Integrin-ligand binding properties govern cell migration speed through cell-substratum adhesiveness. Nature. 1997 Feb 6;385(6616):537–540. [PubMed]
  • Polte TR, Hanks SK. Interaction between focal adhesion kinase and Crk-associated tyrosine kinase substrate p130Cas. Proc Natl Acad Sci U S A. 1995 Nov 7;92(23):10678–10682. [PMC free article] [PubMed]
  • Polte TR, Hanks SK. Complexes of focal adhesion kinase (FAK) and Crk-associated substrate (p130(Cas)) are elevated in cytoskeleton-associated fractions following adhesion and Src transformation. Requirements for Src kinase activity and FAK proline-rich motifs. J Biol Chem. 1997 Feb 28;272(9):5501–5509. [PubMed]
  • Reynolds AB, Roesel DJ, Kanner SB, Parsons JT. Transformation-specific tyrosine phosphorylation of a novel cellular protein in chicken cells expressing oncogenic variants of the avian cellular src gene. Mol Cell Biol. 1989 Feb;9(2):629–638. [PMC free article] [PubMed]
  • Richardson A, Parsons JT. Signal transduction through integrins: a central role for focal adhesion kinase? Bioessays. 1995 Mar;17(3):229–236. [PubMed]
  • Richardson A, Parsons T. A mechanism for regulation of the adhesion-associated proteintyrosine kinase pp125FAK. Nature. 1996 Apr 11;380(6574):538–540. [PubMed]
  • Sabe H, Hata A, Okada M, Nakagawa H, Hanafusa H. Analysis of the binding of the Src homology 2 domain of Csk to tyrosine-phosphorylated proteins in the suppression and mitotic activation of c-Src. Proc Natl Acad Sci U S A. 1994 Apr 26;91(9):3984–3988. [PMC free article] [PubMed]
  • Schaller MD, Borgman CA, Cobb BS, Vines RR, Reynolds AB, Parsons JT. pp125FAK a structurally distinctive protein-tyrosine kinase associated with focal adhesions. Proc Natl Acad Sci U S A. 1992 Jun 1;89(11):5192–5196. [PMC free article] [PubMed]
  • Schaller MD, Borgman CA, Parsons JT. Autonomous expression of a noncatalytic domain of the focal adhesion-associated protein tyrosine kinase pp125FAK. Mol Cell Biol. 1993 Feb;13(2):785–791. [PMC free article] [PubMed]
  • Schaller MD, Hildebrand JD, Shannon JD, Fox JW, Vines RR, Parsons JT. Autophosphorylation of the focal adhesion kinase, pp125FAK, directs SH2-dependent binding of pp60src. Mol Cell Biol. 1994 Mar;14(3):1680–1688. [PMC free article] [PubMed]
  • Schaller MD, Otey CA, Hildebrand JD, Parsons JT. Focal adhesion kinase and paxillin bind to peptides mimicking beta integrin cytoplasmic domains. J Cell Biol. 1995 Sep;130(5):1181–1187. [PMC free article] [PubMed]
  • Schaller MD, Parsons JT. Focal adhesion kinase and associated proteins. Curr Opin Cell Biol. 1994 Oct;6(5):705–710. [PubMed]
  • Schaller MD, Parsons JT. pp125FAK-dependent tyrosine phosphorylation of paxillin creates a high-affinity binding site for Crk. Mol Cell Biol. 1995 May;15(5):2635–2645. [PMC free article] [PubMed]
  • Schlaepfer DD, Hanks SK, Hunter T, van der Geer P. Integrin-mediated signal transduction linked to Ras pathway by GRB2 binding to focal adhesion kinase. Nature. 1994 Dec 22;372(6508):786–791. [PubMed]
  • Schlaepfer DD, Hunter T. Evidence for in vivo phosphorylation of the Grb2 SH2-domain binding site on focal adhesion kinase by Src-family protein-tyrosine kinases. Mol Cell Biol. 1996 Oct;16(10):5623–5633. [PMC free article] [PubMed]
  • Thomas SM, Soriano P, Imamoto A. Specific and redundant roles of Src and Fyn in organizing the cytoskeleton. Nature. 1995 Jul 20;376(6537):267–271. [PubMed]
  • Weiner TM, Liu ET, Craven RJ, Cance WG. Expression of focal adhesion kinase gene and invasive cancer. Lancet. 1993 Oct 23;342(8878):1024–1025. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Cited in Books
    Cited in Books
    PubMed Central articles cited in books
  • Compound
    Compound
    PubChem Compound links
  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...