Differential and Special Properties of the Major Human UGT1-encoded Gastrointestinal UDP-glucuronosyltransferases Enhance Potential to Control Chemical Uptake

Abstract

UDP-glucuronosyltransferase (UGT) isozymes detoxify metabolites, drugs, toxins, and environmental chemicals via conjugation to glucuronic acid. Based on the extended UGT1 locus combined with Northern blot analysis and in situ hybridization, we determined the distribution of UGT1A1 and UGT1A7 through UGT1A10 mRNAs and found them for the first time segmentally distributed in the mucosal epithelia layer of the gastrointestinal tract. Biochemically, recombinant isozymes exhibited pH optima of 5.5, 6.4, 7.6, 8.5, and/or a broad pH range, and activities were found to be unaffected or progressively inhibited by increasing substrate concentrations after attaining V max for certain chemicals. Under different optimal conditions, all exhibited wide substrate selections for dietary and environmentally associated chemicals. Evidence also suggests tandem effects of isozymes in the time for completion of reactions when comparing short-and long-term incubations. Moreover, treatment of colon cells with certain dietassociated constituents, curcumin and nordihydroguaiaretic acid, reversibly targets UGTs causing inhibition without affecting protein levels; there is no direct inhibition of control UGT using curcumin as substrate in the in vitro assay. In summary, we demonstrate that UGTs are located in gastrointestinal mucosa, have vast overlapping activities under differential optimal conditions, and exhibit marked sensitivity to certain dietary substrates/constituents, representing a first comprehensive study of critical properties concerning glucuronidating isozymes in alimentary tissues. Additionally, the highly dynamic, complex, and variable properties necessarily impact absorption of ingested chemicals and therapeutic drugs.The gastrointestinal (GI) 1 system has the critical function of absorbing nutrients from the diet and, simultaneously, acting as a barrier to ingested toxins and unwanted chemicals. To this end, the human UGT1 complex locus, which encodes multiple UDP-glucuronosyltransferase (UGT) isozymes distributed in the GI tract, participates in the broad and critical function of detoxifying lipid-soluble phenols derived metabolically or ingested as part of the diet, its contaminants, and as medications. UGT conjugates glucuronic acid to the acceptor substrate, converting the lipophile to an inactive glucuronide that undergoes facilitated excretion. The isozymes metabolize endogenous substrates such as bilirubin, steroids, bile acids, retinoic acid, and thyroid hormones. Equally important, isozymes also metabolize four categories of dietary phytochemicals that could pose risks for humans and animals when absorbed in excess. Highly abundant dietary flavonoids found in plants are shown to inhibit aromatase (1), and less abundant anthraquinones and two types of phytoestrogens (2-4) can also cause cellular damage. Particular anthraquinones were mutagenic when bioactivated (3) and were carcinogens in long-term feeding experiments in mice (5). Injurious effects of diets were demonstrated by extensive lesions in the reprodu...

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

“…In fact, this expression profile is very similar to the one previously described for other UGTs. 21 The co-localization further supports that exon 5b-containing mRNAs are processed in translational pathways similar to other UGT1As. Moreover, according to the sequence similarity between both i1 and i2 proteins, they are likely to localize in the same subcellular Figure 5 Expression of UGT1As in colon cancer cells transfected with non-target or exon 5b siRNA probes.…”

Targeted inhibition of glucuronidation markedly improves drug efficacy in mice—A model

“…While UGT location in GI mucosa cells [1] suggests it is a strategic site for manipulating glucuronidation by either negative or positive regulators of activities, it is important that evidence for each UGT tested with curcumin [2,3] requires on-going phosphorylation. At appropriate concentrations, the literature indicates chemopreventive curcumin acts to scavenge free radicals and interrupt PKC signaling triggered by oxidants [23].…”

“…Whereas MPA readily forms ether-linked-(MPAG) or acyl-glucuronides (AcMPAG) [9-11], we studied MPA glucuronidation in vitro with human UGTs and found 4 isozymes are avid metabolizers [12] that reach saturation kinetics between 1.6 and 2.4 mM with Km values between 0.25 and 0.55 mM. These UGT isozymes, encoded at the UGT1 complex locus, were found strategically and differentially expressed in the gastrointestinal (GI) mucosa [1]. In this report, we established under both in-vitro and in-vivo conditions that mouse Ugt1a1 also requires phosphorylation, which can be transiently downregulated by nontoxic kinase inhibitor, curcumin [2,3,13], and irreversibly by calphostin-C, similar to that for human UGTs [2,3].…”

“…Because many orally-administered drugs have been shown to undergo glucuronidation under in-vitro conditions with recombinant UGTs, it is possible that GI-glucuronidation has an impact on a significant number of medications leading to premature removal. It remains to be seen, however, whether the significant improvement in both serum free-MPA level and immunosuppression in response to downregulation of GI-distributed glucuronidating activities in mice has human relevance.While UGT location in GI mucosa cells [1] suggests it is a strategic site for manipulating glucuronidation by either negative or positive regulators of activities, it is important that evidence for each UGT tested with curcumin [2,3] requires on-going phosphorylation. At appropriate concentrations, the literature indicates chemopreventive curcumin acts to scavenge free radicals and interrupt PKC signaling triggered by oxidants [23].…”

Epigenetic regulation of the tissue-specific expression of human UDP-glucuronosyltransferase (UGT) 1A10

“…In particular, UGT1A7, UGT1A8, and UGT1A10 are exclusively expressed in the gastrointestinal tract, excluding the liver. This expression limits the bioavailability of orally administered drugs, such as raloxifene, naloxon, and mycophenolic acid, as well as xenobiotics, such as resveratrol and quercetin [5,6]. The intestine-specific expression of UGT1A8 and UGT1A10 was explained by transcriptional regulation through an intestine-specific transcription factor, caudal type homeobox 2 (Cdx2), as well as Sp1 and hepatocyte nuclear factor (HNF) 1α [7,8,9,10].…”