Genetic diversity at the UGT1 locus is amplified by a novel 3′ alternative splicing mechanism leading to nine additional UGT1A proteins that act as regulators of glucuronidation activity

Girard, Hugo; Lévesque, Éric; Bellemare, Judith; Journault, Kim; Caillier, Bertrand; Guillemette, Chantal

doi:10.1097/fpc.0b013e3282f1f118

Cited by 82 publications

(101 citation statements)

References 30 publications

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“…6 These experiments were conducted using two anti-UGT1A antibodies that recognize classical 55 kDa UGT1As_i1 and proteins of 45 kDA corresponding to UGT1A_i2 isoforms. The highest UGT1A_i2 proteins expression was observed in gastrointestinal tissues.…”

Section: Resultsmentioning

confidence: 99%

“…6 All amplification reactions were repeated at least twice in duplicate for both cell lines using independent RT reactions, and positively controlled in tissue expressing UGT of interest. GAPDH was amplified in each RT reaction as a positive RT-PCR control.…”

Section: Characterization Of a New Polyclonal Anti-ugt1a Isoforms 2 Amentioning

confidence: 99%

“…Physiological function of UGT1A_i2 spliced forms J Bellemare et al described 6,7,11,12 and was reported as glucuronidation rates (area or pmol min -1 mg -1 protein).…”

Section: Glucuronide Formation Was Measured By Hplc-ms/ms Asmentioning

confidence: 99%

“…The presence of exon 5b in the mRNA sequence induces the replacement by a 10-amino acid sequence of the residues 435-533 encoded by exon 5a, leading to products with a distinctive C-term feature but retaining the co-substrate-binding domain. 6,7 Further works revealed by the heterologous expression of novel UGT1A_i2 proteins indicate that these spliced forms have undetectable glucuronic acid transferase activity and are therefore enzymatically inactive. Expression studies with microsomes prepared from human tissues using currently available anti-UGT antibodies have highlighted that hepatic but particularly extrahepatic tissues may be a predilection site for those new alternative splicing events.…”

Section: Introductionmentioning

confidence: 99%

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Alternative-splicing forms of the major phase II conjugating UGT1A gene negatively regulate glucuronidation in human carcinoma cell lines

et al. 2009

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The UDP-glucuronosyltransferase UGT1A gene is a major biotransformation gene involved in the metabolism of a vast array of molecules. Recently, we uncovered a new series of alternative spliced isoforms referred to as isoforms 2 or UGT1As_i2 that use an alternative exon 5 (5b). The function of such mRNAs and the corresponding 45 kDa proteins still remains unclear. Although devoid of glucuronosyltransferase activity, UGT1As_i2 are widely co-expressed with the enzymatically active and classical UGT1A isoforms (UGT1As_i1). In this study, we observed abundant signal in human colon tissue samples, predominantly along intestinal crypts. In human cells, UGT1A_i2 proteins are expressed in similar subcellular compartments as UGT1As_i1. Cellular properties of i2-spliced forms were then studied using synthetic small-interfering RNA (siRNA) in two human colon cancer cell lines that show a significant amount of exon 5a-and exon 5b-containing mRNAs and that display enzymatic activities for UGT1As substrates. We observed that siRNA-mediated knockdown of endogenous i2 upregulates cellular glucuronidation activities by 120-170% (Po0.01) for all substrates tested. Functional data support a dominant-negative function for endogenous exon 5b-spliced forms of UGT1A, hence potentially affecting in vivo glucuronidation capacity. This new regulatory strategy may ensure an additional mean to modulate cellular response to endo/xeno stimulus.

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Section: Resultsmentioning

confidence: 99%

Section: Characterization Of a New Polyclonal Anti-ugt1a Isoforms 2 Amentioning

confidence: 99%

“…Physiological function of UGT1A_i2 spliced forms J Bellemare et al described 6,7,11,12 and was reported as glucuronidation rates (area or pmol min -1 mg -1 protein).…”

Section: Glucuronide Formation Was Measured By Hplc-ms/ms Asmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Alternative-splicing forms of the major phase II conjugating UGT1A gene negatively regulate glucuronidation in human carcinoma cell lines

et al. 2009

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“…Heterodimerization has been shown to up-or down-regulate UGT activity [25]. In addition, heterodimers may include recently identified inhibitory UGTs [27]. However, more work is needed to characterize these homo-and hetero-oligomers and their functional implications.…”

Section: Using Mutants and Chimeric Constructs Meech And Mackenzie Dementioning

confidence: 99%

Topological aspects of oligomeric UDP-glucuronosyltransferases in endoplasmic reticulum membranes: Advances and open questions

Bock

Köhle

2009

Biochemical Pharmacology

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This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A c c e p t e d M a n u s c r i p t 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64

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Nuclear Receptor‐Mediated Regulation of Phase II Conjugating Enzymes

Barbier

2008

Nuclear Receptors in Drug Metabolism

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Genetic diversity at the UGT1 locus is amplified by a novel 3′ alternative splicing mechanism leading to nine additional UGT1A proteins that act as regulators of glucuronidation activity

Cited by 82 publications

References 30 publications

Alternative-splicing forms of the major phase II conjugating UGT1A gene negatively regulate glucuronidation in human carcinoma cell lines

Alternative-splicing forms of the major phase II conjugating UGT1A gene negatively regulate glucuronidation in human carcinoma cell lines

Topological aspects of oligomeric UDP-glucuronosyltransferases in endoplasmic reticulum membranes: Advances and open questions

Nuclear Receptor‐Mediated Regulation of Phase II Conjugating Enzymes

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