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Mol Cell Biol. Jun 1996; 16(6): 2606–2613.
PMCID: PMC231251

Introduction of p130cas signaling complex formation upon integrin-mediated cell adhesion: a role for Src family kinases.

Abstract

Integrin-mediated cell adhesion triggers intracellular signaling cascades, including tyrosine phosphorylation of intracellular proteins. Among these are the focal adhesion proteins p130cas (Cas) and focal adhesion kinase (FAK). Here we identify the kinase(s) mediating integrin-induced Cas phosphorylation and characterize protein-protein interactions mediated by phosphorylated Cas. We found that expression of a constitutively active FAK in fibroblasts results in a consecutive tyrosine phosphorylation of Cas. This effect required the autophosphorylation site of FAK, which is a binding site for Src family kinases. Integrin-mediated phosphorylation of Cas was not, however, compromised in fibroblasts lacking FAK. In contrast, adhesion-induced tyrosine phosphorylation of Cas was reduced in cells lacking Src, whereas enhanced phosphorylation of Cas was observed Csk- cells, in which Src kinases are activated. These results suggest that Src kinases are responsible for the integrin-mediated tyrosine phosphorylation of Cas. FAK seems not to be necessary for phosphorylation of Cas, but when autophosphorylated, FAK may recruit Src family kinases to phosphorylate Cas. Cas was found to form complexes with Src homology 2 (SH2) domain-containing signaling molecules, such as the SH2/SH3 adapter protein Crk, following integrin-induced tyrosine phosphorylation. Guanine nucleotide exchange factors C3G and Sos were found in the Cas-Crk complex upon integrin ligand binding. These observations suggest that Cas serves as a docking protein and may transduce signals to downstream signaling pathways following integrin-mediated cell adhesion.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Avraham S, London R, Fu Y, Ota S, Hiregowdara D, Li J, Jiang S, Pasztor LM, White RA, Groopman JE, et al. Identification and characterization of a novel related adhesion focal tyrosine kinase (RAFTK) from megakaryocytes and brain. J Biol Chem. 1995 Nov 17;270(46):27742–27751. [PubMed]
  • Bhattacharya S, Fu C, Bhattacharya J, Greenberg S. Soluble ligands of the alpha v beta 3 integrin mediate enhanced tyrosine phosphorylation of multiple proteins in adherent bovine pulmonary artery endothelial cells. J Biol Chem. 1995 Jul 14;270(28):16781–16787. [PubMed]
  • Birge RB, Fajardo JE, Mayer BJ, Hanafusa H. Tyrosine-phosphorylated epidermal growth factor receptor and cellular p130 provide high affinity binding substrates to analyze Crk-phosphotyrosine-dependent interactions in vitro. J Biol Chem. 1992 May 25;267(15):10588–10595. [PubMed]
  • Birge RB, Fajardo JE, Reichman C, Shoelson SE, Songyang Z, Cantley LC, Hanafusa H. Identification and characterization of a high-affinity interaction between v-Crk and tyrosine-phosphorylated paxillin in CT10-transformed fibroblasts. Mol Cell Biol. 1993 Aug;13(8):4648–4656. [PMC free article] [PubMed]
  • Bockholt SM, Burridge K. Cell spreading on extracellular matrix proteins induces tyrosine phosphorylation of tensin. J Biol Chem. 1993 Jul 15;268(20):14565–14567. [PubMed]
  • Bockholt SM, Burridge K. An examination of focal adhesion formation and tyrosine phosphorylation in fibroblasts isolated from src-, fyn-, and yes- mice. Cell Adhes Commun. 1995 May;3(2):91–100. [PubMed]
  • Burridge K, Turner CE, Romer LH. Tyrosine phosphorylation of paxillin and pp125FAK accompanies cell adhesion to extracellular matrix: a role in cytoskeletal assembly. J Cell Biol. 1992 Nov;119(4):893–903. [PMC free article] [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]
  • Chan PY, Kanner SB, Whitney G, Aruffo A. A transmembrane-anchored chimeric focal adhesion kinase is constitutively activated and phosphorylated at tyrosine residues identical to pp125FAK. J Biol Chem. 1994 Aug 12;269(32):20567–20574. [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]
  • Chen Q, Kinch MS, Lin TH, Burridge K, Juliano RL. Integrin-mediated cell adhesion activates mitogen-activated protein kinases. J Biol Chem. 1994 Oct 28;269(43):26602–26605. [PubMed]
  • Clark EA, Brugge JS. Integrins and signal transduction pathways: the road taken. Science. 1995 Apr 14;268(5208):233–239. [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]
  • Eide BL, Turck CW, Escobedo JA. Identification of Tyr-397 as the primary site of tyrosine phosphorylation and pp60src association in the focal adhesion kinase, pp125FAK. Mol Cell Biol. 1995 May;15(5):2819–2827. [PMC free article] [PubMed]
  • Feller SM, Knudsen B, Hanafusa H. c-Abl kinase regulates the protein binding activity of c-Crk. EMBO J. 1994 May 15;13(10):2341–2351. [PMC free article] [PubMed]
  • Feller SM, Knudsen B, Hanafusa H. Cellular proteins binding to the first Src homology 3 (SH3) domain of the proto-oncogene product c-Crk indicate Crk-specific signaling pathways. Oncogene. 1995 Apr 20;10(8):1465–1473. [PubMed]
  • Feller SM, Ren R, Hanafusa H, Baltimore D. SH2 and SH3 domains as molecular adhesives: the interactions of Crk and Abl. Trends Biochem Sci. 1994 Nov;19(11):453–458. [PubMed]
  • Frech M, John J, Pizon V, Chardin P, Tavitian A, Clark R, McCormick F, Wittinghofer A. Inhibition of GTPase activating protein stimulation of Ras-p21 GTPase by the Krev-1 gene product. Science. 1990 Jul 13;249(4965):169–171. [PubMed]
  • Gale NW, Kaplan S, Lowenstein EJ, Schlessinger J, Bar-Sagi D. Grb2 mediates the EGF-dependent activation of guanine nucleotide exchange on Ras. Nature. 1993 May 6;363(6424):88–92. [PubMed]
  • Gotoh T, Hattori S, Nakamura S, Kitayama H, Noda M, Takai Y, Kaibuchi K, Matsui H, Hatase O, Takahashi H, et al. Identification of Rap1 as a target for the Crk SH3 domain-binding guanine nucleotide-releasing factor C3G. Mol Cell Biol. 1995 Dec;15(12):6746–6753. [PMC free article] [PubMed]
  • Gout I, Dhand R, Hiles ID, Fry MJ, Panayotou G, Das P, Truong O, Totty NF, Hsuan J, Booker GW, et al. The GTPase dynamin binds to and is activated by a subset of SH3 domains. Cell. 1993 Oct 8;75(1):25–36. [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]
  • Hata Y, Kikuchi A, Sasaki T, Schaber MD, Gibbs JB, Takai Y. Inhibition of the ras p21 GTPase-activating protein-stimulated GTPase activity of c-Ha-ras p21 by smg p21 having the same putative effector domain as ras p21s. J Biol Chem. 1990 May 5;265(13):7104–7107. [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]
  • Hirai H, Varmus HE. Site-directed mutagenesis of the SH2- and SH3-coding domains of c-src produces varied phenotypes, including oncogenic activation of p60c-src. Mol Cell Biol. 1990 Apr;10(4):1307–1318. [PMC free article] [PubMed]
  • Hynes RO. Integrins: versatility, modulation, and signaling in cell adhesion. Cell. 1992 Apr 3;69(1):11–25. [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]
  • Imamoto A, Soriano P. Disruption of the csk gene, encoding a negative regulator of Src family tyrosine kinases, leads to neural tube defects and embryonic lethality in mice. Cell. 1993 Jun 18;73(6):1117–1124. [PubMed]
  • Jhun BH, Rose DW, Seely BL, Rameh L, Cantley L, Saltiel AR, Olefsky JM. Microinjection of the SH2 domain of the 85-kilodalton subunit of phosphatidylinositol 3-kinase inhibits insulin-induced DNA synthesis and c-fos expression. Mol Cell Biol. 1994 Nov;14(11):7466–7475. [PMC free article] [PubMed]
  • Juliano RL, Haskill S. Signal transduction from the extracellular matrix. J Cell Biol. 1993 Feb;120(3):577–585. [PMC free article] [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]
  • Kanner SB, Reynolds AB, Wang HC, Vines RR, Parsons JT. The SH2 and SH3 domains of pp60src direct stable association with tyrosine phosphorylated proteins p130 and p110. EMBO J. 1991 Jul;10(7):1689–1698. [PMC free article] [PubMed]
  • Knudsen BS, Feller SM, Hanafusa H. Four proline-rich sequences of the guanine-nucleotide exchange factor C3G bind with unique specificity to the first Src homology 3 domain of Crk. J Biol Chem. 1994 Dec 30;269(52):32781–32787. [PubMed]
  • Lev S, Moreno H, Martinez R, Canoll P, Peles E, Musacchio JM, Plowman GD, Rudy B, Schlessinger J. Protein tyrosine kinase PYK2 involved in Ca(2+)-induced regulation of ion channel and MAP kinase functions. Nature. 1995 Aug 31;376(6543):737–745. [PubMed]
  • Lowenstein EJ, Daly RJ, Batzer AG, Li W, Margolis B, Lammers R, Ullrich A, Skolnik EY, Bar-Sagi D, Schlessinger J. The SH2 and SH3 domain-containing protein GRB2 links receptor tyrosine kinases to ras signaling. Cell. 1992 Aug 7;70(3):431–442. [PubMed]
  • Matsuda M, Hashimoto Y, Muroya K, Hasegawa H, Kurata T, Tanaka S, Nakamura S, Hattori S. CRK protein binds to two guanine nucleotide-releasing proteins for the Ras family and modulates nerve growth factor-induced activation of Ras in PC12 cells. Mol Cell Biol. 1994 Aug;14(8):5495–5500. [PMC free article] [PubMed]
  • Matsuda M, Mayer BJ, Fukui Y, Hanafusa H. Binding of transforming protein, P47gag-crk, to a broad range of phosphotyrosine-containing proteins. Science. 1990 Jun 22;248(4962):1537–1539. [PubMed]
  • Morino N, Mimura T, Hamasaki K, Tobe K, Ueki K, Kikuchi K, Takehara K, Kadowaki T, Yazaki Y, Nojima Y. Matrix/integrin interaction activates the mitogen-activated protein kinase, p44erk-1 and p42erk-2. J Biol Chem. 1995 Jan 6;270(1):269–273. [PubMed]
  • Nada S, Okada M, Aizawa S, Nakagawa H. Identification of major tyrosine-phosphorylated proteins in Csk-deficient cells. Oncogene. 1994 Dec;9(12):3571–3578. [PubMed]
  • Nassar N, Horn G, Herrmann C, Scherer A, McCormick F, Wittinghofer A. The 2.2 A crystal structure of the Ras-binding domain of the serine/threonine kinase c-Raf1 in complex with Rap1A and a GTP analogue. Nature. 1995 Jun 15;375(6532):554–560. [PubMed]
  • Nojima Y, Morino N, Mimura T, Hamasaki K, Furuya H, Sakai R, Sato T, Tachibana K, Morimoto C, Yazaki Y, et al. Integrin-mediated cell adhesion promotes tyrosine phosphorylation of p130Cas, a Src homology 3-containing molecule having multiple Src homology 2-binding motifs. J Biol Chem. 1995 Jun 23;270(25):15398–15402. [PubMed]
  • Ogawa S, Toyoshima H, Kozutsumi H, Hagiwara K, Sakai R, Tanaka T, Hirano N, Mano H, Yazaki Y, Hirai H. The C-terminal SH3 domain of the mouse c-Crk protein negatively regulates tyrosine-phosphorylation of Crk associated p130 in rat 3Y1 cells. Oncogene. 1994 Jun;9(6):1669–1678. [PubMed]
  • Pawson T. Protein modules and signalling networks. Nature. 1995 Feb 16;373(6515):573–580. [PubMed]
  • Petch LA, Bockholt SM, Bouton A, Parsons JT, Burridge K. Adhesion-induced tyrosine phosphorylation of the p130 src substrate. J Cell Sci. 1995 Apr;108(Pt 4):1371–1379. [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]
  • Ren R, Ye ZS, Baltimore D. Abl protein-tyrosine kinase selects the Crk adapter as a substrate using SH3-binding sites. Genes Dev. 1994 Apr 1;8(7):783–795. [PubMed]
  • Ruoslahti E. Integrins. J Clin Invest. 1991 Jan;87(1):1–5. [PMC free article] [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]
  • Sakai R, Iwamatsu A, Hirano N, Ogawa S, Tanaka T, Mano H, Yazaki Y, Hirai H. A novel signaling molecule, p130, forms stable complexes in vivo with v-Crk and v-Src in a tyrosine phosphorylation-dependent manner. EMBO J. 1994 Aug 15;13(16):3748–3756. [PMC free article] [PubMed]
  • Sasaki H, Nagura K, Ishino M, Tobioka H, Kotani K, Sasaki T. Cloning and characterization of cell adhesion kinase beta, a novel protein-tyrosine kinase of the focal adhesion kinase subfamily. J Biol Chem. 1995 Sep 8;270(36):21206–21219. [PubMed]
  • Sasaoka T, Rose DW, Jhun BH, Saltiel AR, Draznin B, Olefsky JM. Evidence for a functional role of Shc proteins in mitogenic signaling induced by insulin, insulin-like growth factor-1, and epidermal growth factor. J Biol Chem. 1994 May 6;269(18):13689–13694. [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. 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]
  • Sells MA, Chernoff J. Epitope-tag vectors for eukaryotic protein production. Gene. 1995 Jan 23;152(2):187–189. [PubMed]
  • Smith DB, Johnson KS. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. [PubMed]
  • Soriano P, Montgomery C, Geske R, Bradley A. Targeted disruption of the c-src proto-oncogene leads to osteopetrosis in mice. Cell. 1991 Feb 22;64(4):693–702. [PubMed]
  • Stein PL, Lee HM, Rich S, Soriano P. pp59fyn mutant mice display differential signaling in thymocytes and peripheral T cells. Cell. 1992 Sep 4;70(5):741–750. [PubMed]
  • Tanaka S, Morishita T, Hashimoto Y, Hattori S, Nakamura S, Shibuya M, Matuoka K, Takenawa T, Kurata T, Nagashima K, et al. C3G, a guanine nucleotide-releasing protein expressed ubiquitously, binds to the Src homology 3 domains of CRK and GRB2/ASH proteins. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3443–3447. [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]
  • Vuori K, Ruoslahti E. Tyrosine phosphorylation of p130Cas and cortactin accompanies integrin-mediated cell adhesion to extracellular matrix. J Biol Chem. 1995 Sep 22;270(38):22259–22262. [PubMed]
  • Xing Z, Chen HC, Nowlen JK, Taylor SJ, Shalloway D, Guan JL. Direct interaction of v-Src with the focal adhesion kinase mediated by the Src SH2 domain. Mol Biol Cell. 1994 Apr;5(4):413–421. [PMC free article] [PubMed]
  • Yoshida Y, Kawata M, Miura Y, Musha T, Sasaki T, Kikuchi A, Takai Y. Microinjection of smg/rap1/Krev-1 p21 into Swiss 3T3 cells induces DNA synthesis and morphological changes. Mol Cell Biol. 1992 Aug;12(8):3407–3414. [PMC free article] [PubMed]
  • Zhu X, Assoian RK. Integrin-dependent activation of MAP kinase: a link to shape-dependent cell proliferation. Mol Biol Cell. 1995 Mar;6(3):273–282. [PMC free article] [PubMed]

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