De novo expression of the alpha 5 beta 1-fibronectin receptor in HT29 colon-cancer cells reduces activity of c-src. Increase of c-src activity by attachment on fibronectin

• Published in: International journal of cancer 1998-03, Vol.76 (1), p.91-98
• Main Authors: Schmidt, R, Streit, M, Kaiser, R, Herzberg, F, Schirner, M, Schramm, K, Kaufmann, C, Henneken, M, Schaefer-Korting, M, Thiel, E, Kreuser, E-D
• Format: Article
• Language: English
• ISSN: 0020-7136
• DOI: 10.1002/(SICI)1097-0215(19980330)76:1<91::AID-IJC15>3.0.CO;2-J

Changes in integrin expression during malignant transformation have been observed in many tumors. Colon-carcinoma cells show reduced expression or even loss of the alpha 5 beta 1 integrin compared to normal or adenoma cells. To determine the significance of absent alpha 5 beta 1 integrin signaling, we transfected the cDNA coding for the alpha 5 integrin sub-unit into the human colon-carcinoma cell line HT29, which constitutively lacks this subunit but does express the beta 1 subunit. We show here that the newly expressed fibronectin receptor alpha 5 beta 1 generates multiple signals, causing marked changes in cytoskeletal arrangements within a few minutes of adhesion to fibronectin. Cells expressing the alpha 5 beta 1 integrin exhibit the formation of actin stress fibers and focal adhesions, as well as the induction of tyrosine phosphorylation of several proteins, within 10 min. We identified the focal adhesion kinase pp125 super(FAK) and the cytoskeletal protein paxillin as major phosphorylation substrates in these cells. These proteins remained hypophosphorylated when alpha 5-negative control cells were plated on fibronectin. The tyrosine kinase pp60 super(c-src), regarded as central in the regulation of cellular proliferation and constitutively over-expressed in HT29 and in colon-carcinoma cells, showed reduced intrinsic kinase activity in unstimulated HT29 alpha 5 cells. In contrast, fibronectin-induced signaling through alpha 5 beta 1 increased pp60 super(c-src) activity. Moreover, immunoprecipitation of pp60 super(c-src) from extracts of HT29 alpha 5 cells cultivated on fibronectin for 20 min revealed complex formation of pp60 super(c-src) and tyrosine-phosphorylated pp125 super(FAK). Our data suggest that de novo expression of the alpha 5 beta 1 integrin in HT29 colon-cancer cells restores signaling via pp125 super(FAK) and pp60 super(c-src). Thus, loss of this receptor during malignant transformation may contribute to tumor-cell autonomy, while reduced activity of pp60 super(c-src) in HT29 alpha 5-cells may participate directly in growth control.