Wnt/beta-catenin signaling regulates cytokine-induced human inducible nitric oxide synthase expression by inhibiting nuclear factor-kappaB activation in cancer cells.
The human inducible nitric oxide synthase (hiNOS) gene is regulated by nuclear factor kappaB (NF-kappaB) and has recently been shown to be a target of the Wnt/beta-catenin pathway. In this study, we tested the hypothesis that Wnt/beta-catenin signaling might regulate cytokine- or tumor necrosis factor alpha (TNFalpha)-induced hiNOS expression through interaction with NF-kappaB. A cytokine mixture of TNFalpha + interleukin (IL)-1beta + IFNgamma induced a 2- to 3-fold increase in hiNOS promoter activity in HCT116 and DLD1 colon cells, but produced a 2-fold decrease in SW480 colon cancer cells. A similar differential activity was seen in liver cancer cells (HepG2, Huh7, and Hep3B). Overexpression of beta-catenin produced a dose-dependent decrease in NF-kappaB reporter activity and decreased cytokine mixture-induced hiNOS promoter activity. Gel shift for TNFalpha-induced hiNOS NF-kappaB activation showed decreased p50 binding and decreased NF-kappaB reporter activity in the beta-catenin-mutant HAbeta18 cells. Conversely, enhanced p50 binding and increased NF-kappaB reporter activity were seen in HAbeta85 cells, which lack beta-catenin signaling. Coimmunoprecipitation confirmed that beta-catenin complexed with both p65 and p50 NF-kappaB proteins. NF-kappaB-dependent Traf1 protein expression also inversely correlated with the level of beta-catenin. Furthermore, SW480 cells stably transformed with wild-type adenomatous polyposis coli showed decreased beta-catenin protein and increased TNFalpha-induced p65 NF-kappaB binding as well as iNOS and Traf1 expression. Finally, beta-catenin inversely correlated with iNOS and Fas expression in vivo in hepatocellular carcinoma tumor samples. Our in vitro and in vivo data show that beta-catenin signaling inversely correlates with cytokine-induced hiNOS and other NF-kappaB-dependent gene expression. These findings underscore the complex role of Wnt/beta-catenin, NF-kappaB, and iNOS signaling in the pathophysiology of inflammation-associated carcinogenesis.
Du
Qiang
Q
Department of Surgery, TE Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania 15213-2582, USA.
Zhang
Xinglu
X
Cardinal
Jon
J
Cao
Zongxian
Z
Guo
Zhong
Z
Shao
Lifang
L
Geller
David A
DA
D025601:Adenomatous Polyposis Coli Protein / Q000096:biosynthesis / Q000235:genetics; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D016207:Cytokines / Q000494:pharmacology*; D004790:Enzyme Induction / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic; D017491:Genes, APC; D006801:Humans; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009369:Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000494:pharmacology; D052247:Nitric Oxide Synthase Type II / Q000096:biosynthesis* / Q000235:genetics; D015398:Signal Transduction; D014158:Transcription, Genetic; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D051153:Wnt Proteins / Q000378:metabolism*; D051176:beta Catenin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D019014:fas Receptor / Q000096:biosynthesis
D025601:Adenomatous Polyposis Coli Protein; C495270:CTNNB1 protein, human; D016207:Cytokines; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D051153:Wnt Proteins; D051176:beta Catenin; D019014:fas Receptor; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II