Dissertation/Thesis Abstract

Nuclear Receptor Regulation of the UDP-Glucuronosyltransferase 1 Locus and the Role of the Pregnane X Receptor as a Tissue-Specific Master Regulator
by Sinnokrak, Erin Janine, Ph.D., University of California, San Diego, 2010, 199; 3548089
Abstract (Summary)

The UDP-glucuronosyltransferases (UGTs) represent the most significant family of Phase II drug metabolism enzymes and assist in the detoxification and removal of endogenous and exogenous compounds from the body. Using UDP-glucuronic acid as a cosubstrate, the UGTs catalyze the transfer of a polar glucuronic acid moiety to a substrate, thereby increasing its size and hydrophilicity, facilitating excretion. The UGTs are subject to regulation by several nuclear receptors (NRs), including the aryl hydrocarbon receptor (AhR), pregnane x receptor (PXR), constitutive androstane receptor (CAR), and peroxisome-proliferators activated receptor (PPAR). Using a Tg-UGT1 mouse line that expresses all nine functional human UGT1A proteins, it was demonstrated that many of the UGT1 genes are regulated by these nuclear receptors in vivo. In ex vivo primary Tg-UGT1 hepatocyte cultures, PPARα was shown to regulate UGT1A1, and promoter analysis experiments were undertaken to identify a PPARα-responsive element on the UGT1A1 promoter. A DR1 element at -3272 was characterized as the PPARα-responsive sequence. UGT1A4 was also shown to be upregulated by the nuclear receptors in vivo. Experiments to characterize nuclear receptor response elements in the UGT1A4 regulatory region resulted in the identification of an AhR-responsive sequence at -3086 and a PPARα-responsive sequence at -366. A putative PXR-responsive element was identified at -212, and although deletion past or mutation of this element resulted in loss of PXR inducibility, basal transcriptional levels increased dramatically, suggesting that in the absence of an agonist, PXR may act through this element to suppress transcription.

Regulation of the UGTs during hormonal events such as development and pregnancy has been well demonstrated in humans and rats, and pregnant Tg- UGT1 mice exhibit dramatic upregulation of UGT1A protein during pregnancy. Although the hormonal changes that occur during the human menstrual or the mouse estrus cycles are much less severe than in pregnancy, UGT1 regulation was shown to occur during estrus. PXR was hypothesized to be the mediator of the hormonal regulation, and experiments were undertaken to examine the contribution of PXR to UGT1 induction. A Tg-UGT1-PXRnull mouse line was generated and was demonstrated to have no functional PXR. It was expected that induction during pregnancy and estrus would be lost in the absence of PXR, however, induction was maintained. Basal levels of UGT1 transcription were elevated in the Tg-UGT1-PXRnull mice, and it was hypothesized that this could be due to the impact of PXR on cross-talk with other nuclear receptors. Induction studies were performed in Tg-UGT1 and Tg-UGT1-PXRnull mice using CAR and AhR agonists. Induction of CAR and AhR control genes was unaffected by the loss of PXR, but basal expression and inducibility of the UGT1 genes were dramatically different in the PXRnull background.

Indexing (document details)
Advisor: Tukey, Robert H.
Commitee: Lawson, Mark, Mellon, Pamela, Shimasaki, Shunichi, Vale, Wylie
School: University of California, San Diego
Department: Biomedical Sciences
School Location: United States -- California
Source: DAI-B 74/05(E), Dissertation Abstracts International
Subjects: Molecular biology
Keywords: Car, Estrus, Metabolism, Ppar, Pregnane X receptors, Pxr, Tissue-specific master regulators, UDP-glucuronosyltransferase, Ugt
Publication Number: 3548089
ISBN: 978-1-267-83288-7
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