RAT DIHYDRODIOL DEHYDROGENASE - COMPLEXITY OF GENE STRUCTURE AND TISSUE-SPECIFIC AND SEXUALLY DIMORPHIC GENE-EXPRESSION

Dihydrodiol dehydrogenase (DD; EC 1.3.1.20) catalyzes a novel pathway of polycyclic aromatic hydrocarbon (PAH) metabolism in which trans-dih ydrodiols (proximate carcinogens) are oxidized to reactive o-quinones which are cytotoxic and genotoxic. In this study, the complementary DN A for rat liver DD was used to examine the structure and regulation of the DD gene. Southern analysis of rat genomic DNA confirmed that DD i s a member of the multigene aldo-keto reductase superfamily. Conservat ive estimates indicate that the rat DD gene is at least 20-25 kilobase s in length. Northern analysis showed that the rat liver transcript wa s 2.4 kilobases whereas the complementary DNA contains an open-reading frame of 966 nucleotides. Primer extension of male and female polyade nylated RNA indicated that the major transcription start sites are onl y 53 and 54 base pairs upstream from the translation start site, confi rming that the RNA has a very long 3'-untranslated region. In male and female tissues, 2.4 kilobase transcripts predominate in liver, small intestine, and lung, which is consistent with a role for the enzyme in PAH metabolism. Transcripts were also detected in male (prostate)- an d female (ovary, mammary gland, and uterus)-specific tissues. In the o vary, two transcripts were observed of 2.4 and 1.4 kilobases in length . Using benzenedihydrodiol as a model substrate for PAH trans-dihydrod iols, highest levels of DD activity were observed in the liver and sma ll intestine of both sexes. Enzyme activity is 2.5-fold higher in the female liver versus the male liver. This sexual dimorphism can be expl ained by increases in the DD mRNA and enzyme protein as measured by do t-blot and immunotitration analyses, respectively. The latter measurem ents indicate that DD represents 1.0% of the soluble protein in female liver but is only 0.5% of the soluble protein in male liver. Hormonal ablation (ovariectomy and hypophysectomy) abolishes the sexual dimorp hism observed in levels of DD mRNA, enzyme protein, and enzyme activit y. Administration of estrogens to males is sufficient to establish the female pattern of gene expression. These data indicate that DD gene e xpression is hormonally regulated, that estrogens exert their effect a t the level of the mRNA, and that aldo-keto reductases involved in PAH metabolism may have their expression regulated by female sex hormones .