In humans, the UGT1 locus encodes 9 functional UGT1A genes. These genes are expressed in a strict tissue specific fashion, with hepatic UGT1A1 expressed at low levels in individuals that inherit the UGT1A1*28 Gilbert's allele. Experiments conducted in transgenic mice expressing the UGT1A1*28 allele as part of the UGT1 locus (Tg-UGT1*28) have demonstrated that expression of all 9 UGT1A proteins closely mimics the differential control that is observed in human tissues. Remarkably, hepatic UGT1A1 expression is depressed as a result of altered expression of the UGT1A1*28 gene in Tg-UGT1*28 mice. Using Tg-UGT1*28 as a model to study expression of the UGT1 locus, the treatment of mice with compounds that active the Ah receptor (AhR), the pregnenalone X-receptor (PXR), the constitutive androstane receptor (CAR), the peroxisome proliferator- activated receptor alpha (PPAR1) and the liver X-receptor (LXR) all profoundly induce various UGT1A genes. PXR and CAR play key roles in many aspects of drug metabolism since they have been described as xenobiotic sensors and are believed to be regulated by a host of steroid and other hormonal activators. We propose that PXR and CAR are regulated in part by the steady-state levels of steroids and hormones. Thus, to fully understand the role of these xenobiotic receptors towards human UGT1A gene expression and human glucuronidation, mouse genetics will be exploited to examine their contribution towards tissue specific and inducible expression of the UGT1 locus. Experiments will be described demonstrating that regulation of the UGT1 locus in Tg-UGT1*28 mice that are Pxr-null and Car-null participate in the constitutive and inducible expression of the UGT1A genes. Since PXR and CAR have been shown to act in concert with many of the same target genes, this application will focus its efforts on defining the role of PXR and CAR in controlling the inducible and tissue specific expression of the UGT1 locus. Experiments conducted in vitro in primary hepatocytes as well as in vivo in UGT1 mice that lack Pxr and/or Car will examine the molecular mechanisms associated with and further link the relationship between xenobiotic receptor expression and the inducible and humoral control of the UGT1 locus. These studies will be coupled with recent technologies employing a combination of reverse genetics and biochemical analysis that have resulted in the functional deletion of the entire Ugt1 locus in mice and the generation of humanized UGT1 (hUGT1*28) mice. Since hUGT1*28 mice are hyperbilirubinemic as a result of the diminished hepatic UGT1A1 gene expression, these mice will be exploited to study the role of PXR and CAR towrds controlling UGT1A1 gene expression and hyperbilirubinemia. To this end, the specific aims of this competitive renewal are 1. Determine the role of PXR and CAR towards regulation of the human UGT1A1 gene, 2. Determine the role of PXR and CAR towards regulation of the UGT1 locus in vitro, 3. Evaluate the role of human PXR and regulation of the human UGT1 locus. 4. Examine the hormonal implications of PXR and CAR directed activation of the UGT1 locus in maternal liver during pregnancy.

Public Health Relevance

The ability to adequately eliminate steroids, bile acids, drugs and environmental toxicants from our cells and tissues by the enzymatic process that leads to the attachment of glucuronic acid to these agents is a crucial step in protecting the organism from a toxic or carcinogenic episode. How human UDP- glucuronosyltransferases (UGTs) are regulated in vivo (in the tissues) is very poorly understood, primarily because the sophisticated tools necessary to develop animal models carrying these human genes have not been developed. This application is the first attempt to investigate how human UGTs are regulated in mice, and to apply this animal to understand the impact of glucuronidation towards drug-drug interactions, drug metabolism and eventually disease.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Xenobiotic and Nutrient Disposition and Action Study Section (XNDA)
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Okita, Richard T
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University of California San Diego
Schools of Medicine
La Jolla
United States
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Fujiwara, Ryoichi; Yoda, Emiko; Tukey, Robert H (2018) Species differences in drug glucuronidation: Humanized UDP-glucuronosyltransferase 1 mice and their application for predicting drug glucuronidation and drug-induced toxicity in humans. Drug Metab Pharmacokinet 33:9-16
Chen, Shujuan; Tukey, Robert H (2018) Humanized UGT1 Mice, Regulation of UGT1A1, and the Role of the Intestinal Tract in Neonatal Hyperbilirubinemia and Breast Milk-Induced Jaundice. Drug Metab Dispos 46:1745-1755
Lu, Wenqi; Rettenmeier, Eva; Paszek, Miles et al. (2017) Crypt Organoid Culture as an in Vitro Model in Drug Metabolism and Cytotoxicity Studies. Drug Metab Dispos 45:748-754
Hirashima, Rika; Itoh, Tomoo; Tukey, Robert H et al. (2017) Prediction of drug-induced liver injury using keratinocytes. J Appl Toxicol 37:863-872
Yoda, Emiko; Paszek, Miles; Konopnicki, Camille et al. (2017) Isothiocyanates induce UGT1A1 in humanized UGT1 mice in a CAR dependent fashion that is highly dependent upon oxidative stress. Sci Rep 7:46489
Chen, Shujuan; Lu, Wenqi; Yueh, Mei-Fei et al. (2017) Intestinal NCoR1, a regulator of epithelial cell maturation, controls neonatal hyperbilirubinemia. Proc Natl Acad Sci U S A 114:E1432-E1440
Fujiwara, Ryoichi; Mitsugi, Ryo; Uemura, Asuka et al. (2017) Severe Neonatal Hyperbilirubinemia in Crigler-Najjar Syndrome Model Mice Can Be Reversed With Zinc Protoporphyrin. Hepatol Commun 1:792-802
Yueh, Mei-Fei; Chen, Shujuan; Nguyen, Nghia et al. (2017) Developmental, Genetic, Dietary, and Xenobiotic Influences on Neonatal Hyperbilirubinemia. Mol Pharmacol 91:545-553
Mitsugi, Ryo; Sumida, Kyohei; Fujie, Yoshiko et al. (2016) Acyl-glucuronide as a Possible Cause of Trovafloxacin-Induced Liver Toxicity: Induction of Chemokine (C-X-C Motif) Ligand 2 by Trovafloxacin Acyl-glucuronide. Biol Pharm Bull 39:1604-1610
Barateiro, Andreia; Chen, Shujuan; Yueh, Mei-Fei et al. (2016) Reduced Myelination and Increased Glia Reactivity Resulting from Severe Neonatal Hyperbilirubinemia. Mol Pharmacol 89:84-93

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