Abstract
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.
Highlights
In the past, we reported induction of c-raf gene expression in preneoplastic foci and tumor nodules of rat liver after feeding the nongenotoxic hepatocarcinogen Aroclor 1254 [12]
We investigated the consequences of aryl hydrocarbon receptor (AhR) activation on c-raf gene expression in cultures of primary hepatocytes and hepatoma cell lines
After normalizing the densitometric scan to the constitutively expressed glyceraldehyde-3-phosphate dehydrogenase gene, a >10-fold increase in c-raf gene expression was observed with the Ah+-responsive clone, whereas the AhR-negative H4IIE cell line did not respond to this treatment
Summary
We reported induction of c-raf gene expression in preneoplastic foci and tumor nodules of rat liver after feeding the nongenotoxic hepatocarcinogen Aroclor 1254 [12]. This mixture of chlorinated biphenyls contains isomers and congeners, some of which are high-affinity ligands of the aryl hydrocarbon receptor (AhR). It is uncertain whether AhR is capable of activating the c-raf promoter [12, 14] This prompted our interest in exploring molecular interactions of AhR with the c-raf gene.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
