REGULATION OF DRUG METABOLISM AND INFLAMMATION BY PREGNANE X RECEPTOR

Abstract

Liver-enriched nuclear receptor (NR) proteins regulate the expression and activity of several pivotal hepatic biochemical pathways including the uptake, metabolism and excretion of cholesterol, bile acids, glucose, and xenobiotic compounds from the body. The pregnane x receptor (PXR, NR1I2) was first identified in 1998 as a member of the NR superfamily. Over the past decade, it has been well established that PXR functions as a master-regulator of xenobiotic- and drug-inducible expression and activity of numerous genes that encode key members of the phase I and phase II metabolic enzymes, as well as several membrane transporter proteins. In this way, activation of PXR serves as the principal defense mechanism defending the body from toxic insult. Similarly, the PXR protein also forms the molecular basis of an important class of drug-drug interactions in the clinical setting. Moreover, ligand-activated PXR negatively regulates inflammatory processes in both liver and intestine. An integrated model is emerging to reveal a key role for the post-translational modification of PXR in the selective suppression of gene expression, and is opening the door to the study of completely new modes of PXR-mediated gene regulation. This dissertation contributes mainly to two key areas of PXR research: (1) Identification a novel PXR target gene- carboxylesterase 6 (Ces6); (2) a study of the SUMOylation and ubiquitination of PXR protein. The results presented in this dissertation were primarily obtained from mouse and cell-culture systems. Data presented here reveal that activation of the inflammatory response modulates the SUMOylation and ubiquitination status of ligand-bound PXR protein. The SUMOylation and ubiquitination of the PXR protein functions to feedback-repress the inflammatory and xenobiotic responses, respectively. Taken together, the data represent a likely mechanism and provides initial molecular details for the connection between the PXR signaling pathway and inflammation. Studies on post-translational modification of PXR indicate how this protein is converted from a positive regulator in drug metabolism into a transcriptional repressor in inflammatory response. Finally, detailed protocols for purification of mammalian proteins necessary to perform in vitro SUMOylation reactions are presented. Taken together, the work presented in this dissertation contributes to understanding the interface between PXR, drug metabolism, and inflammation, which is expected to produce new opportunities for the development of novel therapeutic strategies.