Document Detail


Modulation of the human glucuronosyltransferase UGT1A pathway by splice isoform polypeptides is mediated through protein-protein interactions.
MedLine Citation:
PMID:  19996319     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
This study investigated the molecular mechanisms underlying the regulatory effect of the newly discovered 45-kDa enzymatically inactive UGT1A spliced polypeptides, named isoform i2, upon UGT1A-mediated glucuronidation. Initially, using an inducible system that mimics the relative abundance of isoforms 1 and 2 of UGT1A1 in human tissues, the rates of formation of glucuronides were significantly reduced. We then used a heterologous system constitutively expressing both isoforms i1 and i2 for an in-depth investigation of the presence of spliced i2 on glucuronidation kinetics. UGT1A1, UGT1A7, and UGT1A8 were selected as candidates for these studies. In all cases, co-expression of i1 and i2 in HEK293 cells leads to a significant reduction of the velocity of the glucuronidation reaction without affecting the affinity (K(m) (app)) for all substrates tested and the K(m) for the co-substrate, UDP-glucuronic acid. The data are consistent with a dominant-negative model of inhibition but do not sustain with an UGT1A_i2-mediated inhibition by competitive binding for substrate or the co-substrate. In contrast, the data from the co-immunoprecipitation experiments are indicative of the existence of a mixture homo-oligomeric (i1-i1 or i2-i2) and hetero-oligomeric (i1-i2) complexes in which the i2-i2 and i1-i2 subunits would be inactive. Thus, protein-protein interactions are likely responsible for the inhibition of active UGT1A_i1 by i2 spliced polypeptides. This new regulatory mechanism may alternatively modulate cellular response to endo/xeno stimulus.
Authors:
Judith Bellemare; Mélanie Rouleau; Mario Harvey; Chantal Guillemette
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-12-08
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  285     ISSN:  1083-351X     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2010 Feb 
Date Detail:
Created Date:  2010-02-01     Completed Date:  2010-04-09     Revised Date:  2014-03-28    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  3600-7     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Alternative Splicing*
Blotting, Western
Cell Line
Ecdysone / pharmacology
Glucuronides / metabolism*
Glucuronosyltransferase / chemistry,  genetics,  metabolism*
Humans
Immunoprecipitation
Isoenzymes / chemistry,  genetics,  metabolism
Kinetics
Peptides / genetics,  metabolism*
Protein Binding / drug effects
Protein Multimerization
Signal Transduction
Substrate Specificity
Transfection
Grant Support
ID/Acronym/Agency:
//Canadian Institutes of Health Research
Chemical
Reg. No./Substance:
0/Glucuronides; 0/Isoenzymes; 0/Peptides; 3604-87-3/Ecdysone; EC 2.4.1.-/UGT1A1 enzyme; EC 2.4.1.17/Glucuronosyltransferase; EC 2.4.1.17/UDP-glucuronosyltransferase, UGT1A8; EC 2.4.1.17/UGT1A7 protein, human
Comments/Corrections

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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