The Same Xenobiotic Response Element Is Required for Constitutive and Inducible Expression of the Mammalian Aldehyde Dehydrogenase-3 Gene

Abstract

The mammalian aldehyde dehydrogenase-3 gene (ALDH3) exhibits several aspects of tissue-specific expression. Certain normal tissues, such as the cornea, constitutively expressALDH3at very high levels. Other tissues, such as normal liver, do not expressALDH3.In liver,ALDH3is inducible by polycyclic aromatic hydrocarbon xenobiotics by an Ah-receptor (AhR)-mediated pathway in which a liganded AhR complexes with nuclear ARNT protein, and the complex binds to a xenobiotic response element (XRE) sequence located near −3.0 kb in theALDH35′ flanking region and initiates transcription. We used our recently developed rat corneal epithelium culture model (Boeschet al., J. Biol. Chem.271, 5150–5157, 1996) to study the molecular basis of constitutiveALDH3expression. Transient transfection assays of corneal epithelium using a battery ofALDH35′ flanking region–CAT reporter gene constructs indicate that high constitutiveALDH3expression involves the samecis-acting elements as xenobiotic-inducedALDH3expression in liver. These elements include a strong basal promoter region and the XRE located near −3.0 kb. Western analysis confirms the presence of AhR and ARNT proteins in 3-methylcholanthrene-treated rat liver, as well as ARNT protein in rat corneal epithelium. No AhR protein is found in rat cornea. The −3.0-kbALDH3XRE region contains multiple overlapping transcription factor binding sequences, including consensus sites for AhR, ARNT, HNF1, HNF4, and C/ebp. Electrophoretic mobility shift assays (EMSAs) indicate that constitutive expression ofALDH3in cornea involves binding of ARNT, HNF1, and HNF4 to theALDH3–XRE in an Ah-receptor-independent, ARNT-requiring manner. Transient transfection ofALDH3–CAT reporter gene constructs possessing a mutation in either the ARNT- or HNF4–DNA binding sites of the XRE confirms the functional importance of these sequence motifs in constitutiveALDH3expression.