Cross-talk between aryl hydrocarbon receptor and mitogen-activated protein kinase signaling pathway in liver cancer through c-raf transcriptional regulation.
c-raf is a serine-threonine kinase and a downstream effector of ras signaling. This kinase plays an essential role in cell proliferation, differentiation, and apoptosis. In the past, we reported induction of c-raf gene expression in rat liver cancer on treatment with a mixture of aryl hydrocarbon receptor (AhR) agonists. This prompted our interest in investigating the role of AhR in the transcriptional regulation of c-raf. Initially, we cloned the rat c-raf promoter and sequenced the genomic DNA and cDNA by Southern blotting and capillary electrophoresis. Then, a genetic algorithm was applied to search for putative AhR-binding sites. DNA-binding activity of AhR was confirmed by electromobility shift assay. We also studied c-raf gene expression in rat hepatoma cell lines with functional and/or devoid AhR and in primary human and rat hepatocyte cultures. Overall, we identified five and three AhR-binding sites in the human and rat c-raf gene, respectively. Treatment of hepatocyte cultures with the AhR antagonist resveratrol reduced DNA binding of AhR. Only rat hepatoma cells with functional AhR responded to 1 nmol/L 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment with >10-fold c-raf mRNA induction. Treatment of human and rat hepatocyte cultures with various AhR-activating chemicals resulted in induction of c-raf gene expression, albeit at different levels. Taken collectively, we show AhR to be a master regulator of c-raf and propose cross-talk between AhR and the mitogen-activated protein kinase signaling pathway in chemically induced hepatocarcinogenesis.
Borlak
Jürgen
J
Fraunhofer Institute of Toxicology and Experimental Medicine, Nikolai-Fuchs-Str.1, 30625 Hannover, Germany. borlak@item.fraunhofer.de
Jenke
Hans Stephan
HS
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D020111:Chlorodiphenyl (54% Chlorine) / Q000494:pharmacology; D004147:Dioxins / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology* / Q000235:genetics*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008969:Molecular Sequence Data; D011078:Polychlorinated Biphenyls / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020239:Receptor Cross-Talk / Q000187:drug effects; D018336:Receptors, Aryl Hydrocarbon / Q000819:agonists / Q000378:metabolism*; D016415:Sequence Alignment; D014158:Transcription, Genetic / Q000187:drug effects
D004147:Dioxins; D018336:Receptors, Aryl Hydrocarbon; D020111:Chlorodiphenyl (54% Chlorine); C025647:clophen A50; D011078:Polychlorinated Biphenyls; D019908:Proto-Oncogene Proteins c-raf; C028451:3,4,3',4'-tetrachlorobiphenyl