Dataset Information

Interaction between oxidative stress sensor Nrf2 and xenobiotic-activated aryl hydrocarbon receptor in the regulation of the human phase II detoxifying UDP-glucuronosyltransferase 1A10.

ABSTRACT: The defense against oxidative stress is a critical feature that prevents cellular and DNA damage. UDP-glucuronosyltransferases (UGTs) catalyze the glucuronidation of xenobiotics, mutagens, and reactive metabolites and thus act as indirect antioxidants. Aim of this study was to elucidate the regulation of UGTs expressed in the mucosa of the gastrointestinal tract by xenobiotics and the main mediator of antioxidant defense, Nrf2 (nuclear factor erythroid 2-related factor 2). Xenobiotic (XRE) and antioxidant (ARE) response elements were detected in the promoters of UGT1A8, UGT1A9, and UGT1A10. Reporter gene experiments demonstrated XRE-mediated induction by dioxin in addition to tert-butylhydroquinone (ARE)-mediated induction of UGT1A8 and UGT1A10, which are expressed in extrahepatic tissue in humans in vivo. The responsible XRE and ARE motifs were identified by mutagenesis. Small interfering RNA knockdown, electrophoretic mobility shifts, and supershifts identified a functional interaction of Nrf2 and the aryl hydrocarbon receptor (AhR). Induction of UGT1A8 and UGT1A10 requires Nrf2 and AhR. It proceeds by utilizing XRE- as well as ARE-binding motifs. In summary, we demonstrate the coordinated AhR- and Nrf2-dependent transcriptional regulation of human UGT1As. Cellular protection by glucuronidation is thus inducible by xenobiotics via AhR and by oxidative metabolites via Nrf2 linking glucuronidation to cellular protection and defense against oxidative stress.

SUBMITTER: Kalthoff S

PROVIDER: S-EPMC2825393 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Interaction between oxidative stress sensor Nrf2 and xenobiotic-activated aryl hydrocarbon receptor in the regulation of the human phase II detoxifying UDP-glucuronosyltransferase 1A10.

Kalthoff Sandra S Ehmer Ursula U Freiberg Nicole N Manns Michael P MP Strassburg Christian P CP

The Journal of biological chemistry 20100106 9

The defense against oxidative stress is a critical feature that prevents cellular and DNA damage. UDP-glucuronosyltransferases (UGTs) catalyze the glucuronidation of xenobiotics, mutagens, and reactive metabolites and thus act as indirect antioxidants. Aim of this study was to elucidate the regulation of UGTs expressed in the mucosa of the gastrointestinal tract by xenobiotics and the main mediator of antioxidant defense, Nrf2 (nuclear factor erythroid 2-related factor 2). Xenobiotic (XRE) and a ...[more]

PMID: 20053997

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Project description:The UDP-glucuronosyltransferase (UGT) isozyme system is critical for protecting the body against endogenous and exogenous chemicals by linking glucuronic acid donated by UDP-glucuronic acid to a lipophilic acceptor substrate. UGTs convert metabolites, dietary constituents, and environmental toxicants to highly excretable glucuronides. Because of difficulties associated with purifying endoplasmic reticulum-bound UGTs for structural studies, we carried out homology-based computer modeling to aid analysis. The search found structural homology in Escherichia coli UDP-galactose 4-epimerase. Consistent with predicted similarities involving the common UDP moiety in substrate/inhibitor, UDP-glucose and UDP-hexanol amine caused competitive inhibition by Lineweaver-Burk plots. Among predicted binding sites N292, K314, K315, and K404 in UGT1A10, two informative sets of mutants K314R/Q/A/E/G and K404R/E had null activities or 2.7-fold higher/50% less activity, respectively. Scatchard analysis of binding data of the affinity ligand, 5-azidouridine-[beta- (32)P]diphosphoglucuronic acid, to purified UGT1A10-His or UGT1A7-His revealed high- and low-affinity binding sites. 2-Nitro-5-thiocyanobenzoic acid-digested UGT1A10-His bound with the radiolabeled affinity ligand revealed an 11.3 and 14.3 kDa peptide associated with K314 and K404, respectively, in a discontinuous SDS-PAGE system. Similar treatment of 1A10His-K314A bound with the ligand lacked both peptides; 1A10-HisK404R- and 1A10-HisK404E showed 1.3-fold greater and 50% less label in the 14.3 kDa peptide, respectively, compared to 1A10-His without affecting the 11.3 kDa peptide. Scatchard analysis of binding data of the affinity ligand to 1A10His-K404R and -K404E showed a 6-fold reduction and a large increase in K d, respectively. Our results indicate that K314 and K404 are required UDP-glcA binding sites in 1A10, that K404 controls activity and high-affinity sites, and that K314 and K404 are strictly conserved in 70 aligned UGTs, except for S321, equivalent to K314, in UGT2B15 and 2B17 and I321 in the inactive UGT8, which suggests UGT2B15 and 2B17 contain suboptimal activity. Hence our data strongly support UDP-glcA binding to K314 and K404 in UGT1A10.

Dataset Information

Interaction between oxidative stress sensor Nrf2 and xenobiotic-activated aryl hydrocarbon receptor in the regulation of the human phase II detoxifying UDP-glucuronosyltransferase 1A10.

Publications

Interaction between oxidative stress sensor Nrf2 and xenobiotic-activated aryl hydrocarbon receptor in the regulation of the human phase II detoxifying UDP-glucuronosyltransferase 1A10.

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