Benzyl Isothiocyanate Induces Nuclear Translocation Of Ahr And Its Heterodimerization With Arnt In Hepa1c1c7 Cells

The biotransformation and detoxification of carcinogens, including 2,3,7,8-tetrachlorodibenzo dioxin, benzo [a] pyrene and 3-methylchloranthrene is dependent upon the activity of phase I and II enzymes, particularly CYP1A1 and NQO1. The transcriptional activation of these enzymes is regulated by the aryl hydrocarbon receptor (AhR) and Nuclear Factor-Erythroid 2-Related Factor. These transcription factors reside in the cytoplasm and when activated, shed their respective chaperones and translocate into the nucleus where they interact with nuclear proteins involved with transcription of genes possessing xenobiotic (XRE) and antioxidant (ARE) response elements. We have observed that the phytochemical benzyl isothiocyanate (BIT) induces nuclear translocation of the AhR independently of traditional ligands in Hepa1c1c7 cells suggesting that it might bind the AhR and affect activity of these enzymes as well as the expression of XRE-responsive genes. As a first step in addressing this hypothesis, we investigated whether BIT activates the AhR, causing it to form heterodimers with ARNT in the nuclei of Hepa1 cells. Cells were treated with 5, 10 and 20 µM BIT or DMSO alone. \#946;-Napthoflavone (BNF) was used as a positive control to visualize AhR/ARNT heterodimerization. Using in vitro-activation of Hepa-1 cell lysates as well as nuclear co-immunoprecipitation and mass spectrometry, we observed that AhR/ARNT heterodimerization occurred in the nucleus of Hepa1 cells treated with BIT similar to what was seen in response to BNF. Additionally, Western blot analysis verified that AhR interacted with ARNT in these cells. The results therefore suggest that BIT alone, a know inducer of Phase II enzyme expression, might activate the AhR and facilitate its binding to several genes that regulate cellular function, including genes associated with activation and detoxification of carcinogens. (Support: NIH MBRS/GM028248; RCMI RR033032) Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 3526.