A novel function for the tumor suppressor p16(INK4a): induction of anoikis via upregulation of the alpha(5)beta(1) fibronectin receptor.
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.
Plath
T
T
Medizinische Klinik mit Schwerpunkt Hepatologie und Gastroenterologie, Charité, Campus Virchow-Klinikum, Berlin, Germany.
Detjen
K
K
Welzel
M
M
von Marschall
Z
Z
Murphy
D
D
Schirner
M
M
Wiedenmann
B
B
Rosewicz
S
S
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D051358:Cyclin-Dependent Kinase 4; D019941:Cyclin-Dependent Kinase Inhibitor p16; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D015536:Down-Regulation / Q000502:physiology; D005353:Fibronectins / Q000494:pharmacology; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D007668:Kidney / Q000166:cytology; D008099:Liver / Q000166:cytology; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D010190:Pancreatic Neoplasms; D010641:Phenotype; D011518:Proto-Oncogene Proteins; D017469:Receptors, Fibronectin / Q000378:metabolism*; D012867:Skin / Q000166:cytology; D014162:Transfection; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000502:physiology
D002352:Carrier Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D005353:Fibronectins; D011518:Proto-Oncogene Proteins; D017469:Receptors, Fibronectin; C495900:CDK4 protein, human; C495902:Cdk4 protein, mouse; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases