Mechanism of benzo[a]pyrene-induced Cyp1a-1 gene expression in mouse Hepa 1c1c7 cells: Role of the nuclear 6 s and 4 s proteins☆

Abstract Treatment of wild-type (wt) aryl hydrocarbon (Ah)-responsive mouse Hepa 1c1c7 cells with benzo[ a ]pyrene (B[ a ]P) caused a concentration-dependent induction of ethoxyresorufin O -deethylase (EROD) activity. In contrast, B[ a ]P was inactive as an inducer in Ah nonresponsive class 1 and class 2 mutant cell lines. In parallel experiments, the nuclear fractions from wt cells treated with 10 −7 m [ 3 H]B[ a ]P contained both the 4 s carcinogen binding protein and the 6 s (Ah receptor) complexes, whereas only the 4 s complex was present in the nuclear fraction of the class 2 mutant cells. The results obtained from cotreatment of wt Hepa 1c1c7 cells with 10 −6 or 10 −7 m B[ a ]P and 5 × 10 −7 or 10 −7 m 6-methyl-1,3,8-trichlorodibenzofuran (MCDF) showed that MCDF inhibited the induction of EROD activity and Cyp1a-1 mRNA levels by B[ a ]P. Moreover, using 10 −7 m [ 3 H]B[ a ]P and unlabeled MCDF, it was shown that MCDF not only inhibited the induction response but also caused a concentration-dependent decrease in levels of the nuclear 6 s complex but not the 4 s complex. In contrast, in situ competition studies with unlabeled 10 −6 m] benzo[ ghi ]-perylene (B [ ghi ]P) resulted in the elimination of the nuclear [ 3 H]B[ a ]P 4 s complex (but not the 6 s complex); however, the EROD activity and Cyp1a-1 mRNA levels in cells treated with 10 −7 m B[ a ]P in the presence or absence of 10 −6 m B[ ghi ]P were not significantly different. These results indicate that the 4 s binding protein is not required for the induction of Cyp1a-1 gene expression in Hepa 1c1c7 cells and suggest that B[ a ]P and 2,3,7,8-tetrachlorodibenzo- p -dioxin induce Cyp1a-1 gene expression via a common mechanism which involves binding to the Ah receptor.