Regulation of Sulfotransferase and UDP-Glucuronosyltransferase Gene Expression by the PPARs

During phase II metabolism, a substrate is rendered more hydrophilic through the covalent attachment of an endogenous molecule. The cytosolic sulfotransferase (SULT) and UDP-glucuronosyltransferase (UGT) families of enzymes account for the majority of phase II metabolism in humans and animals. In general, phase II metabolism is considered to be a detoxication process, as sulfate and glucuronide conjugates are more amenable to excretion and elimination than are the parent substrates. However, certain products of phase II metabolism (e.g., unstable sulfate conjugates) are genotoxic. Members of the nuclear receptor superfamily are particularly important regulators of SULT and UGT gene transcription. In metabolically active tissues, increasing evidence supports a major role for lipid-sensing transcription factors, such as peroxisome proliferator-activated receptors (PPARs), in the regulation of rodent and human SULT and UGT gene expression. This review summarizes current information regarding the regulation of these two major classes of phase II metabolizing enzyme by PPARs

DIFFERENTIAL REGULATION OF INDIVIDUAL SULFOTRANSFERASE ISOFORMS BY PHENOBARBITAL IN MALE RAT LIVER

This paper is available online at http://www.dmd.org ABSTRACT: Xenobiotics that induce the cytochromes P450 also produce changes in rat hepatic sulfotransferase (SULT) gene expression. In the present study, male Sprague-Dawley rats were treated for 3 consecutive days with doses of phenobarbital (PB) that induce cytochrome P450 2B1/2 expression. The effects of PB treatment on hepatic aryl SULT (SULT1) and hydroxysteroid SULT (SULT2) mRNA and immunoreactive protein levels and on mRNA expression of individual SULT1 and SULT2 enzyme isoforms were characterized. PB suppressed SULT1A1 mRNA levels, increased the expression of the SULT-Dopa/tyrosine isoform, and did not produce significant changes in SULT1C1 and SULT1E2 mRNA expression. In rats injected with the highest test dose of PB (100 mg/kg), hepatic SULT1A1 mRNA levels were decreased to ϳ42% of control levels and SULT-Dopa/tyrosine mRNA levels were increased to ϳ417% of vehicle-treated control levels. Like the SULT1 subfamily, individual members of the SULT2 gene subfamily were differentially affected by PB treatment. PB (35, 80, and 100 mg/kg) suppressed SULT20/21 mRNA expression to ϳ61, ϳ30, and ϳ41% of vehicle-treated control levels, respectively. In contrast, SULT60 mRNA levels were increased to ϳ162% of control levels and SULT40/41 mRNA levels were increased to ϳ416% of vehicletreated control levels in rats treated with 100 mg/kg PB. These studies support a complex role for PB-mediated effects on the SULT multigene family in rat liver. Because individual SULT1 and SULT2 enzyme isoforms are known to metabolize a variety of potentially toxic substrates, varied responses to PB among members of the SULT multigene family might have important implications for xenobiotic hepatotoxicity

Transcriptional Regulation of Cytosolic Sulfotransferase 1C2 by Intermediates of the Cholesterol Biosynthetic Pathway in Primary Cultured Rat Hepatocytes s

ABSTRACT Cytosolic sulfotransferase 1C2 (SULT1C2) is expressed in the kidney, stomach, and liver of rats; however, the mechanisms regulating expression of this enzyme are not known. We evaluated transcriptional regulation of SULT1C2 by mevalonate (MVA)-derived intermediates in primary cultured rat hepatocytes using several cholesterol synthesis inhibitors. Blocking production of mevalonate with the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor pravastatin (30 mM), reduced SULT1C2 mRNA content by ∼40% whereas the squalene synthase inhibitor squalestatin (SQ1, 0.1 mM), which causes accumulation of nonsterol isoprenoids, increased mRNA content by 4-fold. Treatment with MVA (10 mM) strongly induced SULT1C2 mRNA by 12-fold, and this effect was blocked by inhibiting squalene epoxidase but not by more distal cholesterol inhibitors, indicating the effects of MVA are mediated by postsqualene metabolites. Using rapid amplification of cDNA ends (RACE), we characterized the 59 end of SULT1C2 mRNA and used this information to generate constructs for promoter analysis. SQ1 and MVA increased reporter activity by ∼1.6-and 3-fold, respectively, from a construct beginning 49 base pairs (bp) upstream from the longest 59-RACE product (-3140:-49). Sequence deletions from this construct revealed a hepatocyte nuclear factor 1 (HNF1) element (-2558), and mutation of this element reduced basal (75%) and MVA-induced (30%) reporter activity and attenuated promoter activation following overexpression of HNF1a or 1b. However, the effects of SQ1 were localized to a more proximal promoter region (-281:-49). Collectively, our findings demonstrate that cholesterol biosynthetic intermediates influence SULT1C2 expression in rat primary hepatocytes. Further, HNF1 appears to play an important role in mediating basal and MVA-induced SULT1C2 transcription

Nonsterol Isoprenoids Activate Human Constitutive Androstane Receptor in an Isoform-Selective Manner in Primary Cultured Mouse Hepatocytes

ABSTRACT Our laboratory previously reported that accumulation of nonsterol isoprenoids following treatment with the squalene synthase inhibitor, squalestatin 1 (SQ1) markedly induced cytochrome P450 (CYP)2B1 mRNA and reporter activity in primary cultured rat hepatocytes, which was dependent on activation of the constitutive androstane receptor (CAR). The objective of the current study was to evaluate whether isoprenoids likewise activate murine CAR (mCAR) or one or more isoforms of human CAR (hCAR) produced by alternative splicing (SPTV, hCAR2; APYLT, hCAR3). We found that SQ1 significantly induced Cyp2b10 mRNA (∼3.5-fold) in primary hepatocytes isolated from both CAR-wild-type and humanized CAR transgenic mice, whereas the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor pravastatin had no effect. In the absence of CAR, basal Cyp2b10 mRNA levels were reduced by 28-fold and the effect of SQ1 on Cyp2b10 induction was attenuated. Cotransfection with an expression plasmid for hCAR1, but not hCAR2 or hCAR3, mediated SQ1-induced CYP2B1 and CYP2B6 reporter activation in hepatocytes isolated from CAR-knockout mice. This effect was also observed following treatment with the isoprenoid trans,trans-farnesol. The direct agonist CITCO increased interaction of hCAR1, hCAR2, and hCAR3 with steroid receptor coactivator-1. However, no significant effect on coactivator recruitment was observed with SQ1, suggesting an indirect activation mechanism. Further results from an in vitro ligand binding assay demonstrated that neither farnesol nor other isoprenoids are direct ligands for hCAR1. Collectively, our findings demonstrate that SQ1 activates CYP2B transcriptional responses through farnesol metabolism in an hCAR1-dependent manner. Further, this effect probably occurs through an indirect mechanism

Cholesterol and bile acids regulate xenosensor signaling in drug-mediated induction of cytochromes P450

Cytochromes P450 (CYP) constitute the major enzymatic system for metabolism of xenobiotics. Here we demonstrate that transcriptional activation of CYPs by the drug-sensing nuclear receptors pregnane X receptor, constitutive androstane receptor, and the chicken xenobiotic receptor (CXR) can be modulated by endogenous cholesterol and bile acids. Bile acids induce the chicken drug-activated CYP2H1 via CXR, whereas the hydroxylated metabolites of bile acids and oxysterols inhibit drug induction. The cholesterol-sensing liver X receptor competes with CXR, pregnane X receptor, or constitutive androstane receptor for regulation of drug-responsive enhancers from chicken CYP2H1, human CYP3A4, or human CYP2B6, respectively. Thus, not only cholesterol 7 alpha-hydroxylase (CYP7A1), but also drug-inducible CYPs, are diametrically affected by these receptors. Our findings reveal new insights into the increasingly complex network of nuclear receptors regulating lipid homeostasis and drug metabolism