CAR and PXR: the xenobiotic-sensing receptors

The xenobiotic receptors CAR and PXR constitute two important members of the NR1I nuclear receptor family. They function as sensors of toxic byproducts derived from endogenous metabolism and of exogenous chemicals, in order to enhance their elimination. This unique function of CAR and PXR sets them apart from the steroid hormone receptors. In contrast, the steroid receptors, exemplified by the estrogen receptor (ER) and glucocorticoid receptor (GR), are the sensors that tightly monitor and respond to changes in circulating steroid hormone levels to maintain body homeostasis. This divergence of the chemical- and steroid-sensing functions has evolved to ensure the fidelity of the steroid hormone endocrine regulation while allowing development of metabolic elimination pathways for xenobiotics. The development of the xenobiotic receptors CAR and PXR also reflect the increasing complexity of metabolism in higher organisms, which necessitate novel mechanisms for handling and eliminating metabolic by-products and foreign compounds from the body. The purpose of this review is to discuss similarities and differences between the xenobiotic receptors CAR and PXR with the prototypical steroid hormone receptors ER and GR. Interesting differences in structure explain in part the divergence in function and activation mechanisms of CAR/PXR from ER/GR. In addition, the physiological roles of CAR and PXR will be reviewed, with discussion of interactions of CAR and PXR with endocrine signaling pathways.     

The xenobiotic receptors PXR and CAR in liver physiology,an update

Pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are two nuclear receptors that are well-known for their roles in xenobiotic detoxification by regulating the expression of drug-metabolizing enzymes and transporters. In addition to metabolizing drugs and other xenobiotics, the same enzymes and transporters are also responsible for the production and elimination of numerous endogenous chemicals, or endobiotics. Moreover, both PXR and CAR are highly expressed in the liver. As such, it is conceivable that PXR and CAR have major potentials to affect the pathophysiology of the liver by regulating the homeostasis of endobiotics. In recent years, the physiological functions of PXR and CAR in the liver have been extensively studied. Emerging evidence has suggested the roles of PXR and CAR in energy metabolism, bile acid homeostasis, cell proliferation, to name a few. This review summarizes the recent progress in our understanding of the roles of PXR and CAR in liver physiology.     

P450 gene induction by structurally diverse xenochemicals: central role of nuclear receptors CAR, PXR, and PPAR.

The biochemistry of foreign compound metabolism and the roles played by individual cytochrome P450 (CYP) enzymes in drug metabolism and in the toxification and detoxification of xenochemicals prevalent in the environment are important areas of molecular pharmacology and toxicology that have been widely studied over the past decade. Important advances in our understanding of the mechanisms through which foreign chemicals impact on these P450-dependent metabolic processes have been made during the past 2 years with several key discoveries relating to the mechanisms through which xenochemicals induce the expression of hepatic P450 enzymes. Roles for three "orphan" nuclear receptor superfamily members, designated CAR, PXR, and PPAR, in respectively mediating the induction of hepatic P450s belonging to families CYP2, CYP3, and CYP4 in response to the prototypical inducers phenobarbital (CAR), pregnenolone 16alpha-carbonitrile and rifampicin (PXR), and clofibric acid (PPAR) have now been established. Two other nuclear receptors, designated LXR and FXR, which are respectively activated by oxysterols and bile acids, also play a role in liver P450 expression, in this case regulation of P450 cholesterol 7alpha-hydroxylase, a key enzyme of bile acid biosynthesis. All five P450-regulatory nuclear receptors belong to the same nuclear receptor gene family (family NR1), share a common heterodimerization partner, retinoid X-receptor (RXR), and are subject to cross-talk interactions with other nuclear receptors and with a broad range of other intracellular signaling pathways, including those activated by certain cytokines and growth factors. Endogenous ligands of each of those nuclear receptors have been identified and physiological receptor functions are emerging, leading to the proposal that these receptors may primarily serve to modulate hepatic P450 activity in response to endogenous dietary or hormonal stimuli. Accordingly, P450 induction by xenobiotics may in some cases lead to a perturbation of endogenous regulatory circuits with associated pathophysiological consequences.     

Functional and structural comparison of PXR and CAR

The nuclear receptors pregnane X receptor (PXR, NR1I2) and constitutive active receptor (CAR, NR1I3) have both been proposed to function as xenosensors, but the details of their respective physiological roles are still being elucidated. We have contrasted these two receptors in a variety of experiments including gene expression assays, cell-based ligand profiling assays, and crystallographic/structural modeling analyses. These data highlight key differences between PXR and CAR.     

Inflammation and drug metabolism: NF-kappB and the CAR and PXR xeno-receptors

Decreased drug metabolism, hyperbilirubinemia and intrahepatic cholestasis are frequently observed during inflammation. Additionally, it has long been appreciated that exposure to drug metabolism-inducing xenobiotics can impair immune function. The nuclear receptor CAR (constitutive androstane receptor or NR1I3) and PXR (pregnane X receptor, NR1I2) control phase I (cytochrome P450 2B and 3A), phase II (GSTA, UGT1A1), and transporter (MDR1, SLC21A6, MRP2) genes involved in drugs metabolism, bile acids and bilirubin clearance in response to xenobiotics. It is well known that inflammation, through the activation of NF-kappaB pathway, leads to a decrease of CAR, PXR and RXRalpha expression and the expression of their target genes. In addition, a new study reveals the mutual repression between PXR and NF-kappaB signaling pathways, providing a molecular mechanism linking xenobiotic metabolism and inflammation.     

IgM and its receptors: Structural and functional aspects

This review combines the data obtained before the beginning of the 1990s with results published during the last two decades. The predominant form of the IgM molecule is a closed ring composed of five 7S subunits and a J chain. The new model of spatial structure of the pentamer postulates nonplanar mushroom-shaped form of the molecule with the plane formed by a radially-directed Fab regions and central protruding portion consisting of Cμ4 domains. Up to the year 2000 the only known Fc-receptor for IgM was pIgR. Interaction of IgM with pIgR results in secretory IgM formation, whose functions are poorly studied. The receptor designated as Fcα/μR is able to bind IgM and IgA. It is expressed on lymphocytes, follicular dendritic cells, and macrophages. A receptor binding IgM only named FcμR has also been described. It is expressed on T- and B-lymphocytes. The discovery of new Fc-receptors for IgM requires revision of notions that interactions between humoral reactions involving IgM and the cells of the immune system are mediated exclusively by complement receptors. In the whole organism, apart from IgM induced by immunization, natural antibodies (NA) are present and comprise in adults a considerable part of the circulating IgM. NA are polyreactive, germ-line-encoded, and emerge during embryogenesis without apparent antigenic stimuli. They demonstrate a broad spectrum of antibacterial activity and serve as first line of defense against microbial and viral infections. NA may be regarded as a transitional molecular form from invariable receptors of innate immunity to highly diverse receptors of adaptive immunity. By means of interaction with autoantigens, NA participate in maintenance of immunological tolerance and in clearance of dying cells. At the same time, NA may act as a pathogenic factor in atherosclerotic lesion formation and in development of tissue damage due to ischemia/reperfusion.     

Molecular interactions of dioxins and DLCs with the xenosensors (PXR and CAR): An in silico risk assessment approach

Dioxins and dioxin‐like compounds (DLCs) are known to cause endocrine disruption in humans and animals. Being lipophilic xenobiotic chemicals, they can be easily absorbed into the biological system from the surrounding environments, thereby causing various health dysfunctions. In the present study, a total of 100 dioxins and DLCs were taken, and their binding pattern was assessed with the xenosensors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) in comparison with the corresponding known inhibitors and a well‐studied endocrine disrupting xenobiotic, bisphenol A (BPA). The nuclear receptors CAR and PXR are known to play a significant role in handling potential toxins by coordinating cellular transport and metabolic functions of the same. Among different endocrine‐disrupting chemicals used in the present study, DLCs (PCDFs and PCBs) elicited better interactions in comparison with the parent dioxin (polychlorinated dibenzodioxins) compounds. On comparing D scores of all the compounds against both the receptors, PCDF 8‐hydroxy‐3,4‐dichlorodibenzofuran (8‐OH‐DCDF) and PCB tetrachlorobenzyltoluene (TCBT) exhibited significant molecular interactions against PXR (−7.633 kcal mol⁻¹) and CAR (−8.389 kcal mol⁻¹), respectively. Predominant interactions were found to be H‐bonding, π‐π stacking, hydrophobic, polar, and van der Waals. By contrast, BPA and some natural ligands tested in this study showed lower binding affinities with these receptors than certain DLCs reported herein, ie, certain DLCs might be more toxic than the proven toxic agent, BPA. Such studies play a pivotal role in the risk assessment of exposure to dioxins and DLCs on human health.     

Molecular and functional aspects of parasite invasion

Apicomplexan parasites have evolved an efficient mechanism to gain entry into non-phagocytic cells, hence challenging their hosts by the establishment of infection in immuno-privileged tissues. Gliding motility is a prerequisite for the invasive stage of most apicomplexans, allowing them to migrate across tissues, and actively invade and egress host cells. In the late 1960s, detailed morphological studies revealed that motile apicomplexans share an elaborate architecture comprising a subpellicular cytoskeleton and apical organelles. Since 1993, the development of technologies for transient and stable transfection have provided powerful tools with which to identify gene products associated with these structures and organelles, as well as to understand their functions. In combination with access to several parasite genomes, it is now possible to compare and contrast the strategies and molecular machines that have been selectively designed by distinct life stages within a species, or by different apicomplexan species, to optimize infection.     

Heterogeneity of neuronal nicotinic acetylcholine receptors: Structural and functional aspects

Decay kinetics of the postsynaptic excitatory currents (EPSC), distribution of the antibodies specific to different α-subunits of neuronal nicotinic acetylcholine receptors (nAChR), and the effects of these antibodies on ACh-induced membrane currents were studied in neurons of different autonomic ganglia of rats. It was shown that α3-, α5- and α7-subunits were present in all studied cultured neurons of the rat superior cervical ganglion (SCG), while the α4-subunit was present only in about half of the neurons; this α-subunit distribution differed from that in cultured intracardial neurons of rats. Two nAChR populations were found in rat SCG neurons, and a series of nAChR populations were found in murine superior mesenteric ganglion neurons; they differed in kinetics of their ion channel activity, voltage dependence and the rate of their open channel blockade. The possible functional role of neuronal nAChR heterogeneity is discussed.     

Molecular and functional characterization of adrenomedullin receptors in pufferfish

The receptors for the calcitonin gene-related peptide (CGRP)/adrenomedullin (AM) family peptides were characterized in the mefugu Takifugu obscurus, a euryhaline fugu species very close to Takifugu rubripes, which has as many as five adrenomedullin genes (AM1-5). CGRP and AM share a G protein-coupled core receptor called calcitonin receptor-like receptor (CLR), and the specificity of the CLR is determined by the interaction with receptor activity-modifying proteins (RAMPs). Through database mining, three CLRs (CLR1-3) and five RAMPs (RAMP1-5) were identified, and all of them were cloned by RT-PCR and characterized by functional expression in COS7 cells in every possible combination of CLR-RAMP. The following combinations generated cAMP in response to physiological concentrations of CGRP, AM1 (an ortholog of mammalian AM), AM2, and AM5: CLR1-RAMP1/4 (CGRP), CLR1-RAMP2/3/5 (AM1), CLR2-RAMP2 (AM1), CLR1-RAMP3 (AM2), and CLR1-RAMP3 (AM5). Their expressions were found by Northern blot analysis to be tissue specific and salinity dependent. For example, CLR1-RAMP5 and CLR1-RAMP2 are expressed specifically in the gill and kidney, respectively, suggesting their involvement in osmoregulation. Furthermore, relatively high levels of CLRs and RAMPs were found in the spleen and ovary, suggesting roles in the immune and female reproductive systems. Immunohistochemistry revealed that AM receptors of the following types are expressed in the locations, indicated in brackets, of the mefugu gill and kidney: CLR1-RAMP5 (interlamellar vessels), CLR2-RAMP2 (pillar cells), and CLR1-RAMP2 (apical side of renal proximal tubule cells).     

AMPA和KA受体通道的分子结构及功能特性

近期的克隆研究证实,尽管α－氨基－３－羧基－５－甲基异恶唑－４－丙酸（ＡＭＰＡ）和红藻氨酸（ＫＡ）受体能被相同的激动剂激活,但它们是不同的相互独立的受体复合体。同时,已经证明非Ｎ－甲基－门冬氨酸（ＮＭＤＡ）受体的某些亚型具有Ｃａ＾２＋通透性,这种特性以前认为只有ＮＭＤＡ受体才具有。     

Molecular aspects of functional differences between alcohol and sorbitol dehydrogenases

The amino acid sequence of sheep liver sorbitol dehydrogenase has been fitted to the high-resolution model of the homologous horse liver alcohol dehydrogenase by computer graphics. This has allowed construction of a model of sorbitol dehydrogenase that provides explanations why sorbitol is not a substrate for alcohol dehydrogenase, why ethanol is not a substrate for sorbitol dehydrogenase, and what determines its specificity for polyols. An important feature of the model is that one of the ligands to the active site zinc atom is a glutamic acid residue instead of a cysteine residue, which is the corresponding ligand in the homologous alcohol dehydrogenases. This is one component of the structural change that can be related to the different substrate specificities, showing how altered enzymic activity might be brought about by structural changes of the kind that it is now possible to introduce by site-directed mutagenesis and recombinant DNA techniques.     

Structural and functional aspects of the receptors for platelet-derived growth factor

Platelet-derived growth factor (PDGF) is a 30 kDa dimer of disulfide-bonded A and B chains. Three isoforms of PDGF have been isolated (PDGF-AA, PDGF-AB and PDGF-BB). These bind with different affinities and specificities to two structurally related cell surface receptors, viz. the α-receptor and the β-receptor. The receptors are transmembrane proteins with an intracellular, ligand-stimulatable protein tyrosine kinase domain. Activation of the receptors is intimately associated with receptor dimerization, and available data suggest that PDGF is a divalent ligand such that one molecule of PDGF binds and dimerizes two receptor molecules. Stimulation of PDGF receptors leads to a cascade of cellular events, which have been shown to require an intact receptor tyrosine kinase activity. However, ligand-induced internalization and degradation of the β-receptor occur essentially independent of the receptor kinase activity. Receptor activation leads to the phosphorylation on tyrosine residues of three enzymes, probably by direct phosphorylation: phospholipase C-γ, phosphatidylinositol 3′ kinase and Raf-1. In certain cells, PDGF β-receptor expression is inducible such that cells in normal tissue in vivo do not express receptors; only in inflammatory lesions or when cells are explanted in vitro, are receptors being expressed. Transformation by the v-sis oncogene is mediated by an autocrine PDGF-like growth factor. Although both the α- and β-receptors are structurally related to the v-fms and v-kit oncogenes, it is not known if the PDGF receptors have a transforming potential. In conclusion, the finding of three isoforms of PDGF that interact with two structurally related receptors implies a finely tuned regulatory network, the role of which in cell growth and transformation remains to be clarified.