A novel function for the tumor suppressor p16(INK4a): induction of anoikis via upregulation of the alpha(5)beta(1) fibronectin receptor

• Published in: The Journal of cell biology Vol. 150; no. 6; pp. 1467 - 1478
• Main Authors: Plath, T, Detjen, K, Welzel, M, von Marschall, Z, Murphy, D, Schirner, M, Wiedenmann, B, Rosewicz, S
• Format: Journal Article
• Language: English
• Published: United States 18.09.2000
• Subjects: Animals; Apoptosis - drug effects; Apoptosis - physiology; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Adhesion - drug effects; Cell Adhesion - physiology; Cell Division - physiology; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinases - metabolism; Down-Regulation - physiology; Fibronectins - pharmacology; Flow Cytometry; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor - physiology; Humans; In Vitro Techniques; Kidney - cytology; Liver - cytology; Mice; Mice, Nude; Neoplasm Transplantation; Pancreatic Neoplasms; Phenotype; Proto-Oncogene Proteins; Receptors, Fibronectin - metabolism; Skin - cytology; Transfection; Tumor Cells, Cultured; Up-Regulation - physiology
• ISSN: 0021-9525
• DOI: 10.1083/jcb.150.6.1467

The tumor suppressor gene p16(INK4a) inhibits the kinase activity of the cyclin-dependent kinase 4-6/cyclin D complexes and subsequent phosphorylation of critical substrates necessary for transit through the G1 phase of the cell cycle. Recent studies suggested that control of the G1/S boundary might not be the sole biological function of p16(INK4a). We hypothesized that p16(INK4a) might influence hitherto unknown critical features of a malignant epithelial phenotype, such as anchorage dependence. Here we provide evidence that stable transfection of p16(INK4a) restitutes apoptosis induction upon loss of anchorage (anoikis) in a variety of human cancer cells. Anoikis in p16(INK4a)-transfected cells was evidenced by DNA fragmentation and poly(ADP-ribose) polymerase cleavage upon cultivation on polyhydroxyethylmethacrylate-coated dishes and was associated with suppression of anchorage-independent growth as well as complete loss of tumorigenicity. p16(INK4a)-mediated anoikis was due to selective transcriptional upregulation of the alpha(5) integrin chain of the alpha(5)beta(1) fibronectin receptor as detected by FACS((R)) analysis, immunoprecipitation, Northern blotting, and nuclear run-on assays. Addition of soluble fibronectin and inhibitory alpha(5) antibodies to nonadherent cells completely abolished p16(INK4a)-mediated anoikis, whereas laminin was ineffective. Furthermore, antisense-induced downregulation of the alpha(5) integrin chain in p16(INK4a)-transfected cells restored resistance to anoikis. These data suggest a novel functional interference between a cell cycle-regulating tumor suppressor gene and membrane-bound integrins, thus regulating a hallmark feature of an epithelial transformed phenotype: susceptibility to anoikis.