Mini reviewCAR and PXR: Xenosensors of endocrine disrupters?
Introduction
The pregnane X receptor (PXR, NR1I2; also known as SXR for steroid and xenobiotic sensor) and the constitutive androstane receptor (CAR, NR1I3) are members of the orphan nuclear receptor subfamily. Orphan nuclear receptors share many of the structural features found in the nuclear receptor family, but lack known physiological ligands. Both PXR and CAR are activated by xenobiotics and act as master regulators of phases I through III involved in the detoxification and elimination of steroids, bile acids, and xenobiotics [1], [2], [3]. The purpose of phases I and II enzymes is to transform compounds into more polar forms that can be transported by phase III proteins across membranes for excretion. Detoxification genes induced by PXR and CAR include several phase I cytochrome P450 enzymes (CYPs) [4], [5], [6], [7], phase II enzymes such as uridine diphospho-glucuronosyltransferases (UDPGT), glutathione-S-transferases (GSTs) and sulfotransferases (SULTs) [3], [8], [9], [10], [11], and the phase III transporters such as the multidrug resistance-associated protein 2 (MRP2) and the multidrug resistance protein (MDR1) [12], [13], [14] (Fig. 1).
The PXR is activated by a wide variety of xenobiotics because it has evolved several features, including a small number of polar residues spaced through a smooth, hydrophobic ligand-binding domain [15]. Naturally occurring steroids, such as the pregnanes that are the immediate precursors of progesterone and formed by the side chain cleavage of cholesterol, are the most efficacious activators of PXR [4]. However, since the concentrations of the individual steroids required to activate the PXR are greater than those detected in biological samples, it remains uncertain whether the PXR has a high-affinity ligand or instead functions as a more generalized steroid sensor [4], [16], [17]. The natural or evolutionary PXR ligand might be a metabolite or structure similar to 5β-pregane-3,20-dione based on its IC50 value [18]. Mouse PXR is activated following exposure to pregnenolone 16α-carbonitrile (PCN, a prototypical CYP3A inducer), several pesticides, glucocorticoids and antiglucocorticoids [4], [19]. Human PXR is activated by xenobiotics such as rifampicin, clotrimazole, and hyperforin, and bile acids such as lithocholic acid and 6-keto lithocholic acid [20], [21], [22].
Both activation and inactivation of CAR activity can occur and inactivators are often referred to as inverse agonists since CAR has high constitutive activity in vitro [23]. CAR received its name from the androstanes, which act as inverse agonists. CAR is activated by several drugs, toxicants and steroids, such as phenobarbital, clotrimazole, chlorpromazine, metyrapone, 5β-pregnane-3,20-dione, and 1,4-bis-(2-(3,5-dichloropyridyloxy)) benzene (TCPOBOP) [24], [25], [26], [27]. However, CAR is considered less promiscuous than PXR [28]. Interestingly, ligand binding is not a prerequisite for activation of CAR as demonstrated by phenobarbital [29].
Detoxification genes induced by PXR and CAR include several cytochrome P450 enzymes (CYPs), crucial in the oxidative metabolism of a wide range of endobiotics and xenobiotics. Initially, CAR and PXR were thought to act as regulators of CYP2B and CYP3A, respectively [1], [4], [30]. More recently, experiments have illustrated CAR's ability to activate the CYP3A4 xenobiotic response element, normally thought as the consensus sequence for PXR [24]. This cross-talk is also shown by PXR's ability to activate CYP2B6 by binding to the phenobarbital-responsive enhancer module region of the gene, a region also known to mediate the induction of CYP2B6 by CAR [6], [31], [32], [33]. Other P450s such as CYP2A and CYP2C family members are also induced by PXR and CAR [4], [5], [6], [7], [34].
The purpose of this review is to investigate the role of PXR and CAR as protectors of the endocrine system from chemical perturbation. Tables are provided that show PXR and CAR are activated by a large number of endocrine disrupting chemicals. Properties that allow activation of PXR and CAR by such an extensive number of endocrine disrupters will be discussed. Further, we will consider species differences in PXR and CAR and how this must be taken into account when extrapolating to humans. Lastly, we will provide specific instances where PXR and CAR activation may lead to detrimental effects.
Section snippets
Detoxification enzymes controlled by PXR and CAR
The CYPs are phase I enzymes and hemoproteins that mono-oxygenate, reduce, and hydrolyze various substrates thus making them polar, water-soluble metabolites that can be conjugated by phase II enzymes and removed more rapidly. CYPs are grouped into families, subfamilies and isoforms. The human CYP genes have been arranged into 18 families and 43 subfamilies [35]. CYPs sharing greater than 40% identity at the amino acid level are grouped into the same family with those sharing greater than 55%
Endocrine disrupting and endocrine acting chemicals
Endocrine disrupting chemicals (EDCs) refer to anthropogenic compounds that are able to mimic, antagonize, alter, or modify normal hormonal activity. Endocrine acting chemicals (EACs) encompass EDCs, but also include natural metabolic substances and by-products such as pregnenolone, androstanol, and estrone. Several EACs and EDCs are known to activate PXR and CAR, and the number of EACs that activate these receptors may provide clues as to their roles. This suggests that PXR and CAR are
Activation of PXR and CAR by EACs and EDCs
The human and rodent PXR are activated by a large number of endogenous hormones, their synthetic derivatives and EDCs (Table 2, Table 3). The significant number of EDCs that activate the PXR suggests that it has a role in protecting cells from xenobiotic chemicals that may perturb the endocrine system. That the PXR is also activated by pregnanes and other metabolic products of steroids, further implicates it in the protection of the endocrine system. These chemicals (EACs, EDCs) probably have
Common structural motifs of PXR and CAR activators
Many nuclear receptors, including PXR, CAR, ER, AR, PR, etc., are activated by steroidal ring structures. Just like the estrogen, progesterone, glucocorticoid, and androgen receptors, PXR and CAR received their names from steroidal ligands. The chemicals that activate PXR and CAR vary widely in structure, but many show similarity to the ligands bound by other nuclear receptors in that they have similar molecular weights and are hydrophobic molecules [96]. For example, both ER and PXR are
Species differences
The ligand-binding domain of PXR exhibits a considerable amount of variation among different species. Percent sequence identity of this region in hPXR compared to mPXR and rat PXR is only 77 and 76%, respectively [28]. There are numerous accounts of ligands that activate the human PXR, but not the rodent PXR [81], [102]. For example, rifampicin is a potent activator of hPXR but not of mPXR, while PCN is a weak activator of hPXR and a much more potent activator of mPXR [20]. The chemical
Deleterious consequences of PXR/CAR activation and EAC metabolism
The P450s induced by PXR and CAR are expressed in uninduced livers [3], [28], [108], and involved in the metabolism of steroids and other endogenous compounds [109]. Therefore, testosterone hydroxylation is an often used biomarker of P450 induction [109], [110], [111], [112]. Endogenous hormones, their synthetic derivatives, and EDCs such as the listed pesticides activate PXR (Table 2, Table 3) and sometimes CAR (Table 4, Table 5). In turn, they induce CYP2B, CYP3A and other steroid
Summary
The trend of overlapping ligands bound by both PXR and CAR is exhibited again when looking at EDCs. Further, the ability of PXR and CAR to transcriptionally activate the same consensus sequences and induce many of the same detoxication enzymes provides a level of protection that is redundant, but beneficial at defending diverse organisms from toxicants. The impressive number of EDCs that activate these receptors, especially the PXR, suggests they play a role in protecting the integrity of the
Acknowledgments
Support for this work was provided by NIH grants 2 S06 GM008012-33 and 5G12RR08124.
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