Abstract

Glucocorticoids regulate the rat glutathioneS-transferase A2 (GSTA2) gene in a biphasic manner in cultured hepatocytes that repress gene expression at low concentration (10–100 nM) but induce gene expression at high concentration (>1 μM). High concentrations of the glucocorticoid receptor (GR) antagonist RU38486 (5–10 μM) also induced the expression of GSTA2. These effects were reproduced in HepG2 cells transfected with a luciferase reporter containing 1.6 kilobase pairs of 5′-flanking sequence of GSTA2 and expression plasmids for either GR, pregnane X receptor (PXR) or a combination of both. Dexamethasone t-butylacetate (1 μM t-Bu-DEX) repressed gene expression between 60 to 75% when only GR was expressed. When PXR was expressed, both basal andt-Bu-DEX-dependent gene expression was increased over 2-fold, respectively. Biphasic regulation of gene expression was observed over a broad range of t-Bu-DEX concentrations when expression plasmids for both receptors were cotransfected. Other steroids of the pregnane class induced GSTA2 expression as expected for a PXR-dependent process. Because no canonical responsive element for the PXR-RXRα heterodimer was observed in the 5′-flanking region of the construct, deletion analysis was used to identify a pregnane responsive region between base pairs −700 and −683; this 20-bp region contains the antioxidant response element (ARE). When the ARE sequence was mutated, basal,t-butylhydroquinone- and 17α-hydroxypregnenolone-inducible expression were all lost. These results suggest that PXR interacts with factors binding to the ARE to elicit the pregnane inductive response for GSTA2.