Dexamethasone Regulation of the Rat 3α-Hydroxysteroid/Dihydrodiol Dehydrogenase Gene

Abstract

Rat liver 3α-hydroxysteroid/dihydrodiol dehydrogenase (3α-HSD/DD), a member of the aldo-keto reductase superfamily, inactivates circulating steroid hormones and may contribute to the carcinogenicity of polycyclic aromatic hydrocarbons (PAHs) by oxidizingtrans-dihydrodiols to reactive o-quinones with the concomitant generation of reactive oxygen species. The3α-HSD/DD gene has been cloned, and its 5′-flanking region contains a negative response element (NRE; −797 to −498 bp) that may repress constitutive expression by binding to Oct transcription factors. Upstream from the NRE are three distal imperfect glucocorticoid response elements (GRE1, GRE2, and GRE3); in addition, a proximal imperfect GRE (GRE4) is adjacent to an Oct binding site in the NRE. When rat hepatocytes were cultured on Matrigel and exposed to dexamethasone (Dex), steady state levels of 3α-HSD/DD mRNA were increased 4-fold in a dose-dependent manner, yielding an EC₅₀ value of 10 nm. Time to maximal response was 24 hr, and the effect was blocked with the anti-glucocorticoid RU486. Measurement of the half-life of 3α-HSD/DD mRNA, with and without Dex treatment, indicated that the increase in steady state mRNA levels was not due to increased mRNA stability. By contrast, nuclear run-off experiments using nuclei obtained from Dex-stimulated hepatocytes indicated that Dex increased transcription of the rat3α-HSD/DD gene. Tandem repeats of the imperfect GRE1, GRE2, GRE3, and GRE4 were inserted into thymidine kinase-chloramphenicol acetyl-transferase vectors and cotransfected with the human glucocorticoid receptor into human hepatoma cells. On treatment with Dex, maximal trans-activation of the chloramphenicol acetyl-transferase reporter gene activity was mediated via the proximal GRE (GRE4). These data imply that GRE4 is a functionalcis-element and that binding of the occupied glucocorticoid receptor to this element increases3α-HSD/DD gene transcription. A model is proposed for the positive and negative regulation of the rat3α-HSD/DD gene by the glucocorticoid receptor and Oct transcription factors, respectively.