Protein-DNA interactions at a dioxin-responsive enhancer. Mutational analysis of the DNA-binding site for the liganded Ah receptor.

The liganded Ah receptor activates transcription by binding to a specific DNA-recognition motif within a dioxin-responsive enhancer upstream of the CYP1A1 gene. Analyses of mutant enhancers by gel retardation reveal that each base pair within the domain 5'CGTG(GCAC)3' is essential to the receptor-enhancer interaction. The three base pairs immediately flanking each end of the essential domain contribute less strongly to receptor binding. Analyses of enhancer function by transfection reveal that a mutation in the essential domain, which abolishes receptor-DNA binding, obliterates enhancer function. Mutations outside the essential domain, which diminish, but do not abolish, receptor-DNA binding, also obliterate enhancer function. Additionally, one mutation adjacent to the essential binding motif does not affect receptor-DNA binding, but destroys enhancer activity. These findings imply that transcriptional enhancement by the dioxin-responsive system cannot be predicted solely by the strength of the receptor-enhancer interaction.