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dc.contributor.advisorStaudinger, Jeffrey L
dc.contributor.authorCui, Wenqi
dc.date.accessioned2018-03-09T22:23:07Z
dc.date.available2018-03-09T22:23:07Z
dc.date.issued2017-05-31
dc.date.submitted2017
dc.identifier.otherhttp://dissertations.umi.com/ku:15193
dc.identifier.urihttp://hdl.handle.net/1808/26151
dc.description.abstractPregnane X Receptor (PXR, NR1I2) is a member of the nuclear receptor (NR) superfamily of ligand-activated transcription factors. Expression levels of PXR are highest in the liver and intestine. The activation of PXR can be achieved by exposure to a myriad of xenobiotic compounds and prescription drugs to regulate the expression of genes that encode key enzymes and membrane transporter proteins. Collectively, these PXR-target genes encode gene products that function in a coordinate manner and comprise a vital xenobiotic detoxification pathway in these tissues. In this way, PXR activation by these compounds functions as a ‘xeno-sensor’ of foreign substances in our body to positively regulate the transcription of genes such as cytochrome P450 (CYP) 3A4, and the drug efflux transporter multiple drug resistant protein 1 (MDR1/P-gp), as well as other drug metabolizing enzymes and drug transporter proteins. Xenobiotic-mediated activation of PXR in humans also represents the molecular mechanism of CYP3A4-triggered adverse drug-drug interactions in which the induction of the expression of this broadly selective drug-metabolizing enzyme increases the metabolism of many other co-administered substrates. Activation of PXR also appears to be involved at some level in the MDR1-mediated acquired resistance to chemotherapeutic agents in multiple cancer types. Beyond the canonical physiology of ligand-mediated PXR activation in the regulation of drug metabolism, accumulating evidence clearly indicates that PXR exerts a trans-repressive activity towards the inflammatory response in both the liver and intestine in humans. A broad spectrum of evidence suggests the involvement of post-translational modifications (PTMs) in the regulation of the trans-repressive transcriptional effects of many liver-enriched NR proteins. Previous studies in our laboratory have revealed that PXR is the molecular target of several PTMs including phosphorylation, ubiquitination, and SUMO- (small ubiquitin-like modifier) modification (SUMOylation). Moreover, our research shows that PTMs that target PXR likely regulate its biological activity through sophisticated system of networking or ‘crosstalk’. Crosstalk in this sense is defined as how various PTMs interact with each other on a given protein target to produce a specific biological outcome. The current study is focused on the mechanism of crosstalk between the PTMs and their effect upon the regulation of PXR-mediated trans-repression phenomenon. In the first chapter of this dissertation I provide an introduction to the topic of NR signaling in general, followed by an explanation of canonical PXR signaling in detail. In Chapter 2, the role of crosstalk between the SUMOylation and ubiquitination pathways is examined and its effect upon the regulation of PXR biology in primary hepatocytes is discussed. Tumor necrosis factor-alpha (TNFα)-triggered SUMO(1)ylation of PXR is well-known to inhibit the expression of inflammatory genes in liver and intestine. I show in this dissertation that treatment with the PXR activators, such as Rifampicin (Rif), promotes the SUMO3-modification of PXR. Further, I show that the SUMO(3)ylation of PXR subsequently increases the ubiquitination of PXR, likely to promote proteasomal degradation of this important transcription factor. In Chapter 3, the crosstalk between SUMOylation and acetylation was investigated. I found that pharmacological inhibition of histone deacetylase 3 (HDAC3) activity in cell line-based assays significantly promotes the SUMOylation of PXR, which subsequently impairs the ability of PXR to interact with its canonical corepressor multi-protein complex HDAC3/SMRT (silencing mediator for retinoid or thyroid–hormone receptor). Taken together, the results presented in this dissertation provide novel insight into the likely molecular mechanisms that regulate the clinically observed PXR-mediated trans-repression phenomenon. Specifically, my results suggest that this phenomenon is controlled by an SUMO-acetyl ‘functional switch’ in which PXR acetylation marks PXR as competent for its subsequent SUMOylation, given the correct physiological extracellular condition, namely inflammation. In Chapter 4, the molecular details of the role of phosphorylation in the regulation of PXR-initiated transcription, and its effect upon the interaction with PXR accessory proteins were examined. Utilizing a liquid chromatography tandem mass spectrometry (LC-MS/MS)-based proteomic approach, two phosphorylation sites (T135 and S221) in PXR were identified in primary mouse hepatocytes. Phosphorylation at identified sites inhibits the trans-activation capacity of PXR through interrupting PXR-RXRα hetero-dimerization and PXR association with coactivator proteins. In conclusion, PTMs modulate different aspects of PXR biological activity in the liver and is especially essential for PXR-originated trans-repression of the inflammatory response in liver and intestine. Collectively, the data presented in this dissertation sheds new light upon the molecular mechanisms governing PXR-mediated suppression of inflammation, and could be expected to provide innovative strategies to target the PXR protein for the treatment of inflammatory diseases.
dc.format.extent216 pages
dc.language.isoen
dc.publisherUniversity of Kansas
dc.rightsCopyright held by the author.
dc.subjectPharmacology
dc.subjectInflammation
dc.subjectLiver
dc.subjectPhosphorylation
dc.subjectPregnant x receptor
dc.subjectSUMOylation
dc.subjectUbiquitination
dc.titlePost-translational Modifications of Pregnant X Receptor
dc.typeDissertation
dc.contributor.cmtememberMuma, Nancy
dc.contributor.cmtememberShi, Honglian
dc.contributor.cmtememberAzuma, Yoshiaki
dc.contributor.cmtememberXu, Liang
dc.thesis.degreeDisciplinePharmacology & Toxicology
dc.thesis.degreeLevelPh.D.
dc.identifier.orcid
dc.rights.accessrightsopenAccess


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