Novel Polymorphic Human UDP-glucuronosyltransferase 2A3: Cloning, Functional Characterization of Enzyme Variants, Comparative Tissue Expression, and Gene Induction

Abstract

UDP-glucuronosyltransferases (UGTs) are critical to the detoxification of numerous drugs, environmental pollutants, and endogenous molecules. However, as yet not all of the human UGTs have been cloned and characterized. cDNA clones from the UGT2A3 gene (located on chromosome 4q13) were isolated using pooled human liver RNA. Approximately 10% of clones contained a c.1489AϾG nucleotide substitution, yielding proteins with a residue 497 alanine (UGT2A3.2) instead of a threonine (UGT2A3.1). The allele frequency of this polymorphism (rs13128286) was 0.13 in a European-American population as determined by direct DNA sequencing. Of 81 structurally diverse glucuronidation substrates tested, UGT2A3 expressed by a baculovirus system selectively glucuronidated bile acids, particularly hyodeoxycholic acid at the 6-hydroxy position. Apparent K m values of UGT2A3.1 and UGT2A3.2 for hyodeoxycholic acid 6-glucuronidation were 69 Ϯ 7 and 44 Ϯ 12 M, respectively. Of 29 different extrahepatic tissues evaluated by real-time polymerase chain reaction, UGT2A3 mRNA was most highly expressed in small intestine (160% of liver), colon (78% of liver), and adipose tissue (91% of liver). An in silico scan of the proximal UGT2A3 promoter/5Ј-regulatory region identified transcription factor consensus elements consistent with tissueselective expression in liver (HNF1) and intestine (CXD2), as well as induction by rifampicin (pregnane X receptor). In LS180 human intestinal cells, rifampicin increased UGT2A3 mRNA by more than 4.5-fold compared with vehicle, whereas levels were not significantly affected by the arylhydrocarbon receptor ligand ␤-naphthoflavone. This is the first report establishing UGT2A3 as a functional enzyme, and it represents significant progress toward the goal of having a complete set of recombinant human UGTs for comparative functional analyses.

Identification and Functional Characterization of a Novel UDP-Glucuronosyltransferase 2A1 Splice Variant: Potential Importance in Tobacco-Related Cancer Susceptibility

“…UGT2A3 has been shown to be well expressed in the colon, small intestine, and liver, with activity reported against bile acids (Court et al, 2008). UGT2A2 has been reported to be expressed in the nasal mucosa, with broad substrate selectivity reported against various estrogen and phenylphenol metabolites (Sneitz et al, 2009).…”

The Human UDP-Glucuronosyltransferase UGT2A1 and UGT2A2 Enzymes Are Highly Active in Bile Acid Glucuronidation

“…More recently, UGT2A transcripts were detected in several extrahepatic tissues such as the lung, trachea, larynx, intestine, pancreas, and kidney (Bushey et al, 2011(Bushey et al, , 2013. In addition, UGT2A3 mRNA was also detected in the liver, colon, and adipose tissue (Court et al, 2008). Glucuronidation assays revealed UGT2As as steroid-conjugating enzymes (Itaaho et al, 2008;Sten et al, 2009;Sneitz et al, 2011Sneitz et al, , 2013.…”

“…UGT2A3 only showed activity against simple PAHs such as 1-OH-pyrene and 1-naphthol (Bushey et al, 2013). However, when this enzyme was previously assayed toward a larger battery of potential substrates, glucuronide formation was detected with four bile acids (BAs) (Court et al, 2008).…”

“…The UGT2 family is subdivided into two subfamilies, UGT2A and UGT2B, that contain three and nine enzymes, respectively (Mackenzie et al, 2005). Until recently, UGT2A1 and UGT2A2 were thought to be expressed almost exclusively in the nasal epithelium, and consequently they received less attention than members of the UGT1A and UGT2B subfamilies (Lazard et al, 1991;Jedlitschky et al, 1999;Court et al, 2008). More recently, UGT2A transcripts were detected in several extrahepatic tissues such as the lung, trachea, larynx, intestine, pancreas, and kidney (Bushey et al, 2011(Bushey et al, , 2013.…”

“…Glucuronide conjugates represent up to 10% of the BA circulating pool in healthy volunteers (Trottier et al, 2012), and their formation involves either the 3/6-hydroxyl or 24-carboxyl group of the BA steroid nucleus for the respective formation of ether 3/6G or acyl/ester 24G (Gall et al, 1999;Caron et al, 2006;Trottier et al, 2006Trottier et al, , 2012Court et al, 2008;Verreault et al, 2010). Among the human UGT1A and UGT2B, UGT1A3, UGT2B4, and UGT2B7 have a capacity to convert BAs into BA glucuronides (BA-G) in vitro.…”