2013

http://informahealthcare.com/dmrISSN: 0360-2532 (print), 1097-9883 (electronic)

Drug Metab Rev, 2013; 45(2): 166–172! 2013 Informa Healthcare USA, Inc. DOI: 10.3109/03602532.2012.756012

REVIEW ARTICLE

Epigenetic regulation of pregnane X receptor activity

Yanan Tian

Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, Texas, USA

Abstract

Pregnane X receptor (PXR, NR1I2) is a ligand-dependent nuclear receptor (NR) that functionsas a xenobiotic sensor and effector in coordinately regulating expression of genes of thexenobiotic detoxification network. PXR exerts its transcriptional regulatory functions bydimerization with retinoic X receptor RXR, and PXR-RXR complex binds to specific DNAsequences for regulating gene expression. PXR functions are regulated at the epigenetic levelby chromatin modifications, DNA methylation and noncoding RNA. Chromatin modificationsare carried out, in part, through interaction with coregulator complexes, including steroidcoactivators (SRCs), corepressors (NcoR/SMRT), hepatocyte nuclear factor 4 alpha, proliferatoractivated receptor g coactivator 1 alpha and protein arginine methyltransferase 1. PXR can bemodified by acetylation, phosphorylation and sumoylation, and the promoter of PXR can bemethylated at the ‘‘CpG’’ island. These factors collectively determine the ways in which PXRactivity can be regulated, thereby affecting the magnitude and duration of the PXR-regulateddrug metabolic responses. Most studies of PXR focus on its role as a transcription factor, whichis responsible for the generation of messenger RNA. Recent emerging evidence suggests thatPXR regulates gene expression at both transcriptional and translational levels. This reviewhighlights recent research on the epigenetic mechanisms that are found to be important forthe gene-regulatory activity of PXR and discusses their implications in xenobiotic metabolismand adverse drug responses.

Keywords

Epigenetics, PXR, PRMT1, transcriptional reg-ulation, translational regulation

History

Received 4 October 2012Revised 2 December 2012Accepted 3 December 2012Published online 22 April 2013

Introduction

The majority of xenobiotics/drugs, environmental chemicals,

and many endobiotics are metabolically detoxified through

the detoxification pathways regulated by xenobiotic receptors.

In mammals, the major xenobiotic receptors are pregnane X

receptor (PXR), constitutive androstane receptor (CAR)

and aryl hydrocarbon receptor (AhR). A ligand-dependent

Drosophila ortholog of the human PXR and CAR, DHR96

was identified (King-Jones et al., 2006), suggesting the

evolutionary conservation of the xenobiotic receptor for

dealing with harmful xenobiotics. PXR is a major xenobiotic

receptor with ligands covering a wide range of structurally

diverse compounds from exogenous and endogenous sources,

which include clinical drugs, environmental chemicals and

bile acids (Ihunnah et al., 2011). PXR-regulated cytochrome

P450 (CYP) 3A4 (CYP3A4) is known to play a role in the

metabolization of over 50% of clinical drugs (Kliewer &

Willson, 2002). Many PXR-regulated genes, such as CYP3As,

CYP2Cs, and UGT1As, are well documented for their

importance in drug metabolism and adverse drug responses.

Recent research suggests that PXR is a pleiotropic gene

regulator, and in addition to the well-established roles in

regulating xenobiotic metabolism, it was also found to

regulate physiological and pathophysiological processes,

including cell proliferation, tumorigenesis, inflammatory

responses, cholesterol, and lipid and energy homeostasis

(Ihunnah et al., 2011; Wada et al., 2009; Zhou et al., 2009).

These newly discovered functions of PXR, combined with its

wide range of ligands, make PXR an attractive therapeutic

target for disease treatment (Cheng et al., 2012; Gao & Xie,

2012; Jonker et al., 2012; Li et al., 2012).

Cloning and initial characterization of PXR were reported

in 1998 (Bertilsson et al., 1998; Blumberg et al., 1998;

Lehmann et al., 1998). Identification of this critical xenobi-

otic sensor has facilitated the investigation of the regulatory

mechanism of critical drug-metabolizing enzymes and trans-

porters. A distal regulatory module (�8 kb from the tran-

scription start site; TSS) of the CYP3A4 gene was reported to

be critical for regulation by PXR (Goodwin et al., 1999).

Further characterization of the CYP3A4 for the PXR-binding

sites has identified an additional distal-binding module

(between �10.5 and �11.4 kb from TSS) as a constitutive

liver-enhancer module (Matsumura et al., 2004). These PXR-

binding modules may have functions in differentially regu-

lating gene expression through preferentially interacting with

coregulators, such as hepatocyte nuclear factor 4 alpha

(HNF-4a) (Liu et al., 2008). A recent survey based on

multiple experiments with chromatin immunoprecipitation

(ChIP)-seq and ChIP-chip showed that the nuclear receptor

Address for correspondence: Yanan Tian, Department of VeterinaryPhysiology and Pharmacology, Texas A&M University, MS 4466,College Station, 77845 TX, USA. Fax: 979-845-1921. E-mail:ytian@cvm.tamu.edu

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(NR)-binding sites of the regulatory modules are often located

approximately �10 kb from the TSS sites; therefore, existence

of distant modules for NRs, in fact, is quite common

(Tang et al., 2011). In addition to upstream regulatory

regions, ChIP-seq analysis with mouse liver tissues also

indicates a binding site located in downstream sequences as

well as intergenic regions (Cui et al., 2010).

Epigenome, the center stage for adaptive responses to the

changing environment, gene-environment interaction is a

driving force for the evolution of all organisms and this

interaction requires living organisms be highly adaptable to

the ever-changing environment. Adaptability relies on the

plasticity of the epigenome, which is regulated through

reversible changes in chromatin modifications and DNA

methylation. The term epigenetics here refers to the study of

heritable changes in phenotype or gene expression caused

by mechanisms other than changes in the underlying DNA

sequences, and the epigenome refers to the states of poten-

tially heritable epigenetic changes across the genome. ‘‘The

composition of the epigenome within a given cell is a function

of genetic determinants, lineage, and environment’’

(Bernstein et al., 2007). Although a genome is hard-wired

into primary DNA sequences, and is relatively static, an

epigenome is highly plastic and can be modified by the

developmental stages, cellular environment and signaling

stimuli through DNA methylation, chromatin modifications

and noncoding RNAs, including microRNA (miRNA), which

plays important roles in the regulation of translation. Changes

in epigenone can traverse through cellular mitotic division

and become the basis for the ‘‘epigenetic memory’’, and

when the changes persist through the meiotic phase, then the

changes become transgenerational.

The epigenome of PXR-regulated genes may assume

several different epigenetic states. For example, human

CYP3A4, CYP3A5 and CYP3A7 gene expressions are devel-

opmentally regulated (de Wildt et al., 1999), and in mice, the

Cyp3a16 (neonatal isoform) and Cyp3a11 (adult isoform)

genes were found to be developmentally regulated through

trimethylation of histone H3k27 (repression marker) and

H3K4 (activation marker), although DNA methylation of the

promoter did not seem to be involved (Li et al., 2009).

Epigenetic modulating agents, such as 5-aza-20-deoxycyti-

dine, which inhibits DNA methylation and trichostatin A

(TSA; which inhibits histone deacetylation), altered CYP3A

expression in human liver cell line HepG2 (Dannenberg &

Edenberg, 2006). Nuclear factor kappa B (NF-�B) activation

by inflammatory cytokines can potently suppress human

CYP3A4 expression (Gu et al., 2006), and, reciprocally, PXR

activation suppresses inflammation through interaction with

the NF-�B pathway (Zhou et al., 2006). It was reported

that sumoylation of PXR also plays a role in suppressing

inflammatory responses (Hu et al., 2010). PXR-regulated

genes, such as CYP3A4, in a given tissue or cell type may

become nonresponsive, responsive and sensitized (highly

responsive) through epigenetic regulation. Interestingly, the

levels of PXR and CYP3A4 change as humans age; the

amounts of PXR messenger RNA (mRNA) and proteins

in the liver and intestine reach maximal levels in young adults

(15–38 years old) and then subsequently decrease to less

than half of the maximal levels with age (Miki et al., 2005).

The mechanism for the age-related decreases in PXR

expression is not clear. The reversible methylation of CpG

islands in the PXR promoter has been found to control the

levels of PXR in colon cell lines (Habano et al., 2011); the

role of CpG island methylation for the age-associated change

in PXR level needs further investigation. Understanding the

epigenetic mechanisms of the PXR-regulated drug-metabo-

lism network is important to address pharmacologically

important questions. For example, does previous exposure

to a drug or cellular stimuli (e.g., cytokines and growth

hormones) alter epigenomes so as to affect the responses to

subsequent exposure to the drug? Does epigenetic alterations

of the genome play a role in idiosyncratic drug reaction?

Mode of action for PXR-regulated gene expression

The mode of action of PXR-regulated gene expression has

often been analyzed at the transcriptional level with its

prototypic target gene, CYP3A4, an abundant adult human

liver CYP, and its gene expression is highly inducible by the

typical agonist, rifampicin (RIF). Although PXR belongs to

the NR family, it is located in both the cytoplasm and nucleus

and the ligand-activation of PXR facilitates its nuclear

translocation (Squires et al., 2004). Transcriptional regulation

by PXR can be divided and analyzed in three interconnected,

sequential phases: initiation, elongation and termination.

Initiation

PXR-regulated transcription requires dimerization of PXR

with RXR, and the PXR-RXR dimer binds to the conserved

sequences arranged in different configurations. Agonist

binding strengthens the binding between PXR and DNA.

For many NRs, agonist binding changes the confirmation of

the NR to reduce the affinity for corepressor while gaining

the affinity for coactivator. However, a recent in vitro study

suggests that PXR affinity for the corepressor and -activator is

independent of the ligand, RIF, suggesting that the mecha-

nism of activation in PXR differs significantly from those

observed in many other NRs (Navaratnarajah et al., 2012).

Although PXR binds to both the proximal promoter and

distal enhancer module, full induction of CYP3A4 transcrip-

tion requires both proximal and distal modules. The distal

enhancer module can interact with the proximal promoter

through interaction between the receptor and coregulator/

mediator (illustrated in Figure 1). Looping of the DNA strand

brings the distant enhancer module with its associated factors

to the close proximity of the promoter for gene regulation,

leading to the formation of the preinitiation complex with

RNA polymerase II assembled at the promoter; therefore, it is

an important point of regulation. Disruption of this commu-

nication between the enhancer module and promoter

suppresses gene expression (Kodama et al., 2011). The loop

formation can be detected and analyzed by cross-linking the

protein-DNA complex, followed by creating sticky ends by

restriction enzymes and then ligation, followed by polymerase

chain reaction amplification (Tolhuis et al., 2002).

Elongation and termination

Research on transcriptional regulation in the past has been

mainly focused on formation of the preinitiation complex at

DOI: 10.3109/03602532.2012.756012 Epigenetic regulation of PXR activity 167

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the proximal promoter region. However, transcription elon-

gation is not a constant forward process. It is currently

recognized that transcription elongation by RNA Pol II is a

stage that consists of discrete biochemical steps that are

important targets for regulation (Adelman & Lis, 2012). After

assembly of the preinitiation complex for RNA synthesis

at the promoter, transcription elongation by RNA Pol II

is subjected to negative regulation, and the Pol II CTD

(C-terminal domain) is hypophosphorylated, which only

generates short RNA transcripts (‘‘abortive transcription’’),

a phenomenon called ‘‘proximal promoter pausing’’. A recent

survey using ChIP-seq technology found that many promoters

demonstrated enriched RNA Pol II, suggesting that pausing is

a common phenomenon further supporting the notion that

transcription elongation is an important step for regulation

(Levine, 2011; Muse et al., 2007). For the xenobiotic receptor,

AhR, we have found that it regulates cyp1a1 expression at

transcription elongation through interaction with the elonga-

tion factor, P-TEFb, and RNA Pol II can be ‘‘paused’’ at the

cyp1a1 promoter (Tian et al., 2003). It is conceivable that

under certain conditions, the RNA Pol II pausing would

result in its ‘‘piling up’’ (poised) state for much more efficient

transcription when the transcriptional arrest is released. For

drug-inducible genes, such as those regulated by PXR, RNA

Pol II pausing may lead to much stronger responses, which

may cause adverse drug responses.

Transcription termination for protein-coding genes is

typically functionally coupled with an RNA maturation

event in which the 30 end of the nascent transcript undergoes

cleavage followed by polyadenylation. This 30-end processing

reaction can be broken down into two steps: First transcrip-

tion of a poly(A) site is followed by pausing of Pol II

transcription and endoribonucleolytic cleavage of the nascent

transcript; and second, the upstream cleavage product is

polyadenylated, whereas the downstream cleavage product

is degraded (Kuehner et al., 2011). Both elongation and

termination of transcrption are highly regulated involving

multiple regulatory protein complexes. Very little is known

about how these processes are regulated for PXR. These two

regulatory steps are involved in generating polymorphic

RNA transcripts through alternative splicing and alternative

polyadenylation for mRNA. Future investigation of these

critical regulatory mechanisms is highly warranted.

PXR transcriptional activity regulated bycoregulators and –factors

PXR binds to specific DNA sequences consisting of AG[G/

T]TCA or AGAACA half-site motifs in a variety of config-

urations, including direct repeats DR-3, DR-4, and DR-5 and

everted repeats ER-6 and ER-8 in the target genes (Orans

et al., 2005). These motif configurations and their various

positions are ‘‘hard-wired’’ in the genome and interact

dynamically with transcription factors and coregulators in

determining the outcomes of PXR-regulated gene expression.

Various NR coregulators have been reported to interact with

PXR, including steroid receptor coactivator-1 (SRC-1)

(Kliewer et al., 1998), p300/CBP (Xie et al., 2001), RIP140

(Masuyama et al., 2001), proliferator activated receptor gcoactivator 1 alpha (PGC-1a) (Bhalla et al., 2004), HNF-4a(Tirona et al., 2003) and protein arginine methyltransferase 1

(PRMT1) (Xie et al., 2009). In addition to modulating

chromatin remodeling, these interactions are also important

mechanisms for cross-talk between PXR and other pathways.

The cofactors and -regulators may be grouped in a regulatory

hierarchical manner according to their roles in controlling

PXR-regulated tissue/cell-type specific responses.

Recent results of a genome-wide survey using ChIP-seq

and RNA-seq technology with 25 transcription factors suggest

a sequential and temporal binding of transcription factors to

the regulatory regions of genes in dendritic cells in response

to lipopolysaccharide treatment. According to the sequences

Bgl II-9743

-6035

XREM

RXRPXR

Bgl II Bgl II6035

PXR

CDK7

Y S P T S P S

P TEFb

CDK7

Pol II

RXRPXR

P-TEFb

+6433551302

ER 6 TATA +1

Bgl IIBgl II+643-355-1302

Bgl II

RXR

Figure 1. Looping model of the coregulator/cofactor-mediated interaction between distal xenobiotic enhancer module and proximal promoter ofCYP3A4. Ligand-activated PXR/RXR complex binds to both XREM and proximal promoter regions through binding to consensus DNA sequences.XREM and the promoter are brought to the proximity of the promoter region by the coregulators and -factors where RNA Pol II is recruited. Therecruited RNA Pol II needs to be phosphorylated to proceed to the processive elongation phase.

168 Y. Tian Drug Metab Rev, 2013; 45(2): 166–172

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of association with genes, the binding of these factors can

be divided in three groups, including those of a ‘‘pioneer’’,

which are preestablished during the cellular differentiation,

a second group of the transcription factors that bind DNA in a

the unstimulated state and are highly correlated with future

stimulus-dependent gene induction, and, finally, the third

group of genes binding dynamically in a stimulus-dependent

manner (Garber et al., 2012).

In an analogous manner, the cofactors and -regulators of

PXR may be grouped according to their functions during cell

lineage establishment and roles in transcriptional regulation.

It is conceivable that liver-enriched master regulator HNF-4aand master regulator for gut morphogenesis CDX2 (Beck &

Stringer, 2010) are the pioneer coregulators and are

preassociated with the responsive genes. It has been shown

that HNF-4a and CDX2 are both essential for PXR-regulated

responses (Dou et al., 2012; Tirona et al., 2003). In HeLa

cells, transfection of HNF-4a makes it more ‘‘hepatocyte

like’’ in PXR-regulated responses (Tirona et al., 2003). Other

coregulators may be signal dependent for their interactions

with the PXR/RXR complex in the context of epigenome.

Hepatocyte nuclear factor 4 alpha

HNF-4a (NR2A1) is required for the development of the liver

and is a master regulator for NRs (Hwang-Verslues & Sladek,

2010). HNF-4a is an obligatory factor for the gene-regulatory

activity of PXR and CAR (Tirona et al., 2003). HNF-4a binds

to the conserved DNA sequences as a homodimer to direct

repeat elements upstream of the target gene. Deletion of

the HNF-4a-binding site in the CYP3A4 enhancer region

abolishes the ability of PXR and CAR to activate the CYP3A4

promoter. In Caco and HeLa cells, coexpression of PXR and

HNF-4a drastically increase the transaction of the CYP3A4

promoter by the PXR ligand, RIF (Tirona et al., 2003).

Interestingly, although HNF-4a potentiates the transactivation

function of PXR and CAR for CYP3A4 gene expression, PXR

interaction with HNF-4a may lead to suppression, as shown in

sulfotransferase (SULT)1E1 gene regulation. In the transacti-

vation of SULT1E1, HNF-4a binds to the enhancer and acts

as a mediator that enables the SULT1E1 promoter to form an

active chromatin structure by placing the enhancer close to

the proximal promoter. Ligand-activated PXR interacts with

HNF-4a and prevents promoter-enhancer interaction, thereby

repressing transcription of the SULT1E1 gene (Kodama et al.,

2011). Ligand-activated PXR specifically inhibits HNF-4a-

mediated transactivation of CYP7A1 (Bhalla et al., 2004).

Therefore, HNF-4a plays a critical role in controlling the

PXR-regulated metabolic network through distinct mecha-

nisms leading to either activation or repression.

Protein arginine methyltransferase 1

Protein methylation and demethylation plays an important

role in gene expression regulated by NRs (reviewed in Wu &

Zhang, 2009). Methylation is catalyzed by the PRMT family

of enzymes, which utilize intracellular S-adenosyl methionine

as the methyl donor. In mammals, there are nine protein

arginine methyltransferases (PRMT1–PRMT9, PRMT4 also

known as CARM1), which are categorized into four groups

based on the enzyme reaction mechanism and endproducts

(Di Lorenzo & Bedford, 2011). Thus far, there are at least

five arginine residues (H3R2, H3R8, H3R17, H3R26 and

H4R3) and six lysine residues (H3K4, H3K9, H3K27,

H3K36, H3K79 and H4K20) on histones H3 and H4 that

can be methylated (Klose & Zhang, 2007).

The transcriptional activity of PXR is regulated by

PRMT1. PRMT1 directly associates with PXR and plays an

important role in PXR-regulated gene expression. The ChIP

assay showed that PRMT1 is recruited to the regulatory

region of CYP3A4, with a concomitant methylation of

arginine 3 of histone H4, in response to the PXR agonist,

RIF. In mammalian cells, small interfering RNA knockdown

and gene deletion of PRMT1 greatly diminished the tran-

scriptional activity of PXR, suggesting an indispensable role

of PRMT1 in PXR-regulated gene expression. Interestingly,

PXR appears to have a reciprocal effect on the PRMT1

functions by regulating its cellular compartmentalization as

well as its substrate specificity (Xie et al., 2009).

Interestingly, there seems to be a dynamic interaction on

the epigenome between PRMT1 methylation and histone

acetylation. It has been shown that histone modification by

PRMT1 facilitates subsequent histone acetylation (Huang

et al., 2005), forming an interplay between H4R3 methylation

and other histone modifications. For example, arginine

methylation (H4R3) by PRMT1 facilitates H4 acetylation,

but H4 acetylation inhibits methylation of H4R3 (Wang et al.,

2001). These observations suggest that histone modifications

during transcription proceed in an unidirectional sequence,

and to complete a transcription cycle, methylated H4R3 has to

be demethylated, followed by acetylation and then deacetyla-

tion or replacement (Tian, 2009). Understanding of these

mechanistic processes has an interesting implication in

PXR-regulated drug metabolism and drug-drug interaction.

For example, a ‘‘poised’’ or sensitized ‘‘state’’ for PXR-

regulated responses may be established by epigenome-

regulating compounds (which may not be PXR ligands)

through inhibiting histone acetylation and/or promoting

PRMT1-regulated methylation. A much more potent PXR-

regulated response may be generated in the epigenetically

sensitized cells and tissues upon ligand challenge.

The consequence of histone modifications has been

hypothesized to establish the ‘‘histone code’’ (Jenuwein &

Allis, 2001; Strahl & Allis, 2000) that needs to be interpreted

for gene regulation. The ‘‘code reader’’ for PRMT1 methyl-

ated histone has been identified. Recently, Yang et al.

discovered that TDRD3, a Tudor domain protein, functions

as an arginine-methylated histone reader, which preferentially

recognizes H3R12me2a and H4R3me2a marks associated

with the promoter (Yang et al., 2010). Although TDRD3 itself

has no enzymatic activity, it has been speculated that it

functions as a coactivator through interaction with other

as-yet-unidentified proteins for transcripitonal activation

(Yang et al., 2010).

Epigenomes can be manipulated to achieve more-

efficacious results in the treatment of diseases (Law et al.,

2012; Ramaswamy et al., 2012). Because the epigenome can

be sensitized [e.g., by a histone deacetylation inhibitor],

understanding the role of the histone modification enzymes

in NR-regulated gene expression is also important to avoid

adverse drug interaction.

DOI: 10.3109/03602532.2012.756012 Epigenetic regulation of PXR activity 169

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Post-translational modification of PXR

An important aspect of PXR-regulated gene expression has

been revealed recently by analyzing the post-translational

modifications of PXR protein. The characterized post-

translational modifications (PTMs) are ubiquitylated

(Masuyama et al., 2002), phosphorylation (Lichti-Kaiser

et al., 2009a), sumoylation and acetylation (Biswas et al.,

2011; reviewed in Staudinger et al., 2011). Although PRMT1

interacts with PXR to regulate gene expression, and the

PXR-associated PRMT1 complex methylates histone proteins

in vitro, PRMT1 does not directly methylate PXR in vitro

(Xie et al., 2009).

Results of analysis of PTM of PXR suggest that PTM plays

an important role in regulating the transcriptonal activity as

well as cross-talk between different signaling pathways. It was

shown that phosphorylation status of threonine residues was

altered after activation of PKA signaling in cultured hepato-

cytes, and phosphorylation of human PXR caused recruitment

of corepressor multiprotein complexes, thereby producing

repression of drug-inducible PXR target gene expression

(Lichti-Kaiser et al., 2009b). Sumoylation of PXR plays an

important role in repression of the inflammatory pathway in

liver cells through an interesting potential mechanism

whereby activation of the inflammatory response in hepato-

cytes strongly modulates the sumoylation of ligand-bound

PXR and sumoylated PXR contains SUMO3 chains, and

feedback represses the immune responses in hepatocytes by

targeting the liganded PXR to the regulatory regions bound by

inflammatory mediators, such as NF-kB (Hu et al., 2010).

Although the mechanistic details remain to be investigated,

the binding of sumoylated PXR may prevent the dissociation

of corepressor complex from the NF-kB-bound inflammatory

gene, thereby repressing gene expression.

Translational regulation of PXR-controlled geneexpression

The steady-state level of an mRNA is balanced dynamically

by its genesis and degradation. Most studies regarding

molecular functions of PXR focus on its role as a transcription

factor, which is responsible for the generation of mRNA.

Because prolonged activation of the metabolic detoxification

system by PXR is harmful to the organism, it is important that

mRNA turnover and mRNA translation be regulated and

mRNA degraded once the stimulating agonist is eliminiated.

However, hitherto mechanistic understandings are very lim-

ited regarding how mRNA degradation is regulated and

whether PXR plays a direct role in the regulation. Recent

results indicated that PXR (Takagi et al., 2008) and PXR-

regulated CYP3A4 (Pan et al., 2009) are regulated at the

translational level by miRNAs miR-148 a and miR-27 b,

respectively. miRNA regulates gene expression by targeting at

the 30UTR (untranslated region) of mRNA and regulating

mRNA translational efficiency and mRNA stability (Bartel,

2009). Pan et al. showed that the broadly conserved miR-27b

targets the 30UTR of both CYP3A4 and vitamin D receptor

(VDR), leading to negative regulation of CYP3A4 in LS-180

and PNAC1 cells (Pan et al., 2009).

A key step in mRNA degradation in the cytoplasm is the

shortening of the poly(A) tail, which involves several

deadenylase enzymes. A major deadenylation complex is

the CCR4-NOT complex, which consists of nine subunits

(Doidge et al., 2012). The major members of the core CCR4-

NOT complex (Not1-5, Ccr4 and Caf1) were identified in

yeast by genetic selections for their transcriptional regula-

tions. Two subunits in the complex, CCR4 and CAF1, play a

role as deadenylases. CCR4-NOT is a conserved complex

from yeast to humans in regulating gene expression, both

transcriptionally and post-transcriptionally. It has been

reported to suppress NR transcriptional activity (Winkler

et al., 2006). The CCR4-NOT complex was demonstrated to

deadenylate the mRNA associated with RNA-induced silenc-

ing complex in miRNA-accelerated RNA decay (Behm-

Ansmant et al., 2006). Interestingly, we have identified that

PXR directly interacted with CCR4-NOT complex in a yeast

two-hybrid screening, and ongoing studies indicate that

ligand-activated PXR modulates deadenylation activity

(unpublished results). Although a detailed mechanism still

awaits the further research, emerging evidence suggests that

the PXR-controlled xenobiotic metabolic pathway is regulated

Coregulators

XenobioticMetabolism/Disposition

Phase I, II, III Reactions

Transcriptional regulation

CCR4-NOT

Translational regulation

Xenobiotics

Figure 2. An integrated model for the coordinated transcriptional and translational regulation of gene expression by PXR. Ligand-activated PXR/RXRcomplex interacts with the coregulators and chromatin modification enzymes, such as PRMT1, SRC1,2 and P300/CBP for transcriptional regulation,which generates mRNA. In the cytoplasm, PXR interacts with the CCR4-NOT complex and regulates its deadenylase activity, which determines thestability of mRNA for PXR-regulated genes. The molecular mechanism remains to be investigated.

170 Y. Tian Drug Metab Rev, 2013; 45(2): 166–172

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at the translation, as well as transcription, level, and a model

is proposed to suggest that an integrated regulation of gene

expression by PXR at both the transcription and translation

level (Figure 2).

Summary and future directions

In the past decade, PXR and its regulated metabolic

detoxification network have been a rapidly developing area

in pharmacology and toxicology research. Accumulating

evidence suggests that the functions of PXR are regulated

by genetic/epigenetic mechanisms at both the transcription

and translation level. A model integrating both transcriptional

and translational regulation by PXR is schematically illus-

trated in Figure 2, suggesting that a steady-state level of

transcriptome is achieved through the balancing act of RNA

genesis and destruction, both of which are actively regulated

by PXR The increased understandings of the genomic/

epigenomic regulation of gene expression have provided

novel druggable targets, which are the regulatory components

of eipgenome for disease treatment. Many druggable targets

for these types of ‘‘epidrugs’’ are also critical regulators for

xenobiotic receptors, such as PXR, CAR, and AhR. These

pose new challenges for the application of drugs, because the

epigenome-regulating drugs will also alter PXR-regulated

drug metabolism. Further investigation of the epigenetic

mechanisms underpinning PXR-regulated gene expression

will provide important insights for safe drug application and

effective treatment of diseases, including diseases treated

through the targeting of PXR-regulated pathways.

Declaration of Interest

The research was supported, in part, by the National Institute

of Environmental Health Sciences (grant no.: ES09859).

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