Abstract
UDP-glucuronosyltransferase 1A1 (UGT1A1) plays an important physiological role by contributing to the metabolism of endogenous substances such as bilirubin in addition to xenobiotics and drugs. The UGT1A1 gene has been shown to be inducible by nuclear receptors steroid xenobiotic receptor (SXR) and the constitutive active receptor, CAR. In this report, we show that in human hepatoma HepG2 cells the UGT1A1 gene is also inducible with aryl hydrocarbon receptor (Ah receptor) ligands such as 2,3,7,8-tetrachlodibenzo-p-dioxin (TCDD), beta-naphthoflavone, and benzo[a]pyrene metabolites. Induction was monitored by increases in protein and catalytic activity as well as UGT1A1 mRNA. To examine the molecular interactions that control UGT1A1 expression, the gene was characterized and induction by Ah receptor ligands was regionalized to bases -3338 to -3287. Nucleotide sequence analysis of this UGT1A1 enhancer region revealed a xenobiotic response element (XRE) at -3381/-3299. The dependence of the XRE on UGT1A1-luciferase activity was demonstrated by a loss of Ah receptor ligand inducibility when the XRE core region (CACGCA) was deleted or mutated. Gel mobility shift analysis confirmed that TCDD induction of nuclear proteins specifically bound to the UGT1A1-XRE, and competition experiments with Ah receptor and Arnt antibodies demonstrated that the nuclear protein was the Ah receptor. These observations reveal that the Ah receptor is involved in human UGT1A1 induction.
Highlights
Glucuronidation has evolved in vertebrates to catalyze the transfer of glucuronic acid from uridine 5Ј-diphosphoglucuronic acid to soluble non-lipid dependent substances generated as byproducts of dietary and cellular metabolism [1]
We show that in human hepatoma HepG2 cells the UDP-glucuronosyltransferase 1A1 (UGT1A1) gene is inducible with aryl hydrocarbon receptor (Ah receptor) ligands such as 2,3,7,8-tetrachlodibenzo-p-dioxin (TCDD), -naphthoflavone, and benzo[a]pyrene metabolites
It was observed that induction of UGT1A1 RNA and 17␣-ethynylestradiol glucuronidation by TCDD and BNF correlated with increased levels of UGT1A1 protein (Fig. 1C), with BNF generating slightly greater levels of induced UGT1A1 in microsomes
Summary
Glucuronidation has evolved in vertebrates to catalyze the transfer of glucuronic acid from uridine 5Ј-diphosphoglucuronic acid to soluble non-lipid dependent substances generated as byproducts of dietary and cellular metabolism [1]. Human UGT1A1 has recently been shown to be under control by agents that induce gene expression in concordance with the constitutive active receptor (CAR) [9] and the steroid xenobiotic receptor (SXR) [33]. Induction of UGT1A1 by TCDD and BNF—The small phenolic compound 1-naphthol was used as a substrate to examine UGT activity in HepG2 cells (Fig. 1A).
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