The long-term goal of this project is to understand the molecular mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) action. TCDD is a widespread, persistent environmental contaminant that, in animals, produces morphological, immunologic, biochemical, teratogenic, and neoplastic effects. The risk that TCDD poses to humans is unknown, with potential reproductive and carcinogenic effects being of most concern. The induction of CYP1A1 gene transcription in mouse hepatoma cells constitutes a useful model response for analyzing the mechanism of dioxin action. The increase in gene expression requires the binding of TCDD to an intracellular protein (the Ah receptor), followed by the binding of the liganded receptor to a transcriptional enhancer, situated upstream of the target CYP1A1 gene. The experiments proposed here are designed to analyze the receptor-enhancer mechanism in greater depth. The studies involve the use of molecular biological techniques to (a) analyze the structure and function of the dioxin-responsive enhancer, (b) purify the Ah receptor protein and identify its cDNA, and (c) analyze the TCDD-inducible, Ah receptor-dependent protein-DNA interactions at the dioxin-responsive enhancer within the intact cell.
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