Regulation of the rat glutathione S-transferase A2 gene by glucocorticoids: Involvement of both the glucocorticoid and pregnane X receptors

Glucocorticoids regulate the rat glutathione S-transferase A2 (GSTA2) gene in a biphasic manner in cultured hepatocytes that repress gene expression a t low concentration (10-100 nM) but induce gene expression at high concentr ation (>1 muM). High concentrations of the glucocorticoid receptor (GR) ant agonist RU38486 (5-10 muM) also induced the expression of GSTA2. These effe cts were reproduced in HepG2 cells transfected with a luciferase reporter c ontaining 1.6 kilobase pairs of 5'-flanking sequence of GSTA2 and expressio n plasmids for either GR, pregnane X receptor (PXR) or a combination of bot h. Dexamethasone t-butylacetate (1 muM t-Bu-DEX) repressed gene expression between 60 to 75% when only GR was expressed. When PXR was expressed, both basal and t-Bu-DEX-dependent gene expression was increased over 2-fold, res pectively. Biphasic regulation of gene expression was observed over a broad range of t-Bu-DEX concentrations when expression plasmids for both recepto rs were cotransfected. Other steroids of the pregnane class induced GSTA2 e xpression as expected for a PXR-dependent process. Because no canonical res ponsive element for the PXR-RXR alpha heterodimer was observed in the 5'-fl anking region of the construct, deletion analysis was used to identify a pr egnane responsive region between base pairs -700 and -683; this 20-bp regio n contains the antioxidant response element (ARE). When the ARE sequence wa s mutated, basal, t-butylhydroquinone- and 17 alpha -hydroxypregnenolone-in ducible expression were all lost. These results suggest that PXR interacts with factors binding to the ARE to elicit the pregnane inductive response f or GSTA2.