ABSTRACT
Some forms of cytochrome P450 (CYP) genes are known to be induced by xenobiotics such as dioxins. Induction of the CYP1A gene is mediated by an aryl hydrocarbon receptor (AHR) which binds to a specific nucleotide sequence called a dioxin-responsive element (DRE) located in the 5' enhancer region of the gene. Functional analysis of the regulatory region of the eel CYP1A gene had shown that a 654-bp region near the basal promoter, containing no DREs but three motifs that resemble estrogen-responsive element (ERE) halfsites, contributes substantially to the induced expression. Considering the importance of non-DRE elements in CYP1A gene induction, we investigated the role of ERE-like motifs using a point mutation technique. The regulatory region of the eel CYP1A gene was identified by creating mutations in all three ERE half-sites simultaneously or individually. The regulatory region constructs were cloned upstream of the luciferase gene in an expression vector which was microinjected into medaka ova. Expression of luciferase as a foreign gene in medaka fry exposed to 3-methylcholanthrene (3-MC)-treated feed was measured by competitive PCR analysis. Mutation in all the three ERE half-sites reduced expression to 10%. Mutation in ERE(-2) or ERE(-3) reduced the expression to 50%, whereas mutation in ERE(-1) did not affect the induction. The two functional half-sites of the eel CYP1A gene are palindromic to each other and appear to constitute the full-ERE.
