Mutations affecting the regulation of transcription of the cytochrome P1-450 gene in the mouse Hepa-1 cell line.

Abstract

The cytochrome P1-450-dependent activity, aryl hydrocarbon hydroxylase, and cytochrome P1-450 messenger RNA levels were studied in wild-type Hepa1c1c7 cells and in aryl hydrocarbon hydroxylase-deficient mutants derived from this line. Tetrachlorodibenzo-p-dioxin (TCDD) induced both parameters approximately 50-fold in Hepa1c1c7 cells. Mutants in genes B and C that are affected in the functioning of the Ah receptor required for hydroxylase induction and dominant mutants had nondetectable or much reduced P1-450 mRNA levels and hydroxylase activities after TCDD treatment. Hybrids between wild-type cells and the dominant mutants were also deficient in these parameters. The dominant mutants therefore appear to express a trans-acting repressor of cytochrome P1-450 transcription. Mutants in gene A were heterogenous. Some lacked the mRNA completely; others were inducible for it; while still others (subgroup IV) had high levels even when they were grown without TCDD. These results suggest strongly that gene A is the structural gene for cytochrome P1-450. When subgroup IV mutants were cultured together with wild-type cells, they failed to express P1-450 mRNA in the absence of TCDD treatment. These cells probably accumulate an inducer that can be metabolized by aryl hydrocarbon hydroxylase. The inducer did not appear to be a component of the medium and therefore may be an endogenous ligand of the Ah receptor.