Abstract
The Pregnane X Receptor (PXR) is regarded as a central factor in the body's response to alterations in chemical levels: It is activated by a variety of chemically unrelated endogenous and exogenous compounds and regulates genes involved in the uptake, metabolism, and excretion of these chemicals. The aim of this study was to delineate the molecular mechanisms underlying regulation of the PXR gene. Initial in silico analysis of 2200 bp of DNA 5' to the PXR putative transcription start site suggests a complex promoter with binding sites for a large number of both liver-enriched and ligand-activated transcription factors. This region of the PXR proximal promoter was cloned from genomic DNA and a reporter gene assay deletion series constructed. Over-expression of liver-enriched and ligand-activated transcription factors in Huh7 hepatoma cells was used to examine the role of these factors in regulating the PXR reporter gene. The liver-enriched transcription factors HNF3α/β and HNF4α showed positive regulation of fragments containing putative interaction sites for these factors. Amongst the ligand-activated transcription factors examined, GRα, ER and PPARα produced a net positive effect on PXR reporter gene expression, whereas PXR and CAR produced a net inhibitory effect, suggesting feedback regulation of PXR expression by PXR and CAR proteins, potentially to prevent disruption of cellular homeostasis. The effect of PPARα action of PXR gene expression was examined in more detail, and I have demonstrated for the first time that PPARα regulates gene expression of the PXR by binding directly to its promoter. This is of particular interest as PPARα ligands are not classical PXR ligands, suggesting a complex relationship with implications for drug-drug interactions. In addition, PPARα-mediated activation of PXR gene expression is probably a cross species event, in contrast to the rodent-specific carcinogenesis elicited by PPARα ligands, raising the important question of the implications of this activation in man.