FUNCTIONAL ANALYSIS OF NUCLEAR RECEPTOR VARIANTS

Summary

Principal Investigator: Curtis J Omiecinski
Abstract: DESCRIPTION (provided by applicant): The constitutive androstane receptor (CAR, NR1I3) is a member of the nuclear receptor superfamily that is expressed primarily in the liver. CAR plays a key role in regulating all three phases of biotransformation, pathways that determine the fates of metabolism and toxicity associated with exposures to drugs, other xenobiotics, and endogenous substances such as bile acids, thyroid hormone, heme, and steroids. Due to the importance of this receptor system as a determinant of xenobiotic disposition in man, together with CAR's increasingly recognized role as a physiological regulator, it is imperative to fully characterize the biological processes directed by the human CAR proteome. Progress in the previous research program included the discovery of novel forms of CAR, CAR2 and CAR3;receptors generated through the use of alternative splicing mechanisms that result in the insertion of 4- and 5- amino acids, respectively, within the receptors'ligand binding domain. These receptors are expressed simultaneously with CAR in human livers, but possess highly unique functional attributes. Unlike CAR itself, the variant CARs are ligand-activated receptors, exhibit selective chemical activation profiles, and appear to differentially modulate target gene expression. We hypothesize that alternative splicing enhances the functional diversity of CAR molecules, resulting in overlapping, yet discriminating roles as xenobiotic sensors, driving the interplay of hepatic gene expression networks that in turn, provide critical integration signals directing human physiological responses to the chemical milieu. To test our hypotheses, a progression of three specific aims will be deployed that include: 1) the structural analysis of crystallized CAR proteins;2) use of primary human hepatocyte culture models to enable receptor-driven transcriptional profiling, target gene interactions, and assessment of the dynamics of receptor intracellular localization;and, 3) humanized transgenic mouse investigations to analyze the transcriptional controls modulated by the receptors in vivo, and the phenotypic impact of receptor expression within a series of serum markers. We predict that the respective modulation of genomic programs and the resulting signaling circuitry controlled by the variant CARs have critical functional implications for human health, determining outcomes such as drug-drug interactions, drug and bile acid induced hepatotoxicity, carcinogen and steroid metabolism, and the regulation of lipid and energy homeostasis. The results of the proposed research will contribute important new advances and insights regarding the role of the CAR nuclear receptors as direct modulators of the dynamic gene regulatory networks that determine toxicological and physiological responses to both xenobiotic and endogenous substances. PUBLIC HEALTH RELEVANCE: In this research program, our studies will characterize the roles of a novel series of receptor proteins that interact with pharmaceuticals and environmental chemicals and function to regulate the liver's capacity to metabolize substances. These receptors are termed constitutive androstane receptors and are part of a battery of sensing proteins that exist with liver cells to assist in processing the nature of the cell's chemical environment so that it can better tune its ability to detoxify chemicals we are exposed to.
Funding Period: 2002-09-01 - 2014-11-30
more information: NIH RePORT

Top Publications

  1. pmc CAR2 displays unique ligand binding and RXRalpha heterodimerization characteristics
    Scott S Auerbach
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Drug Metab Dispos 35:428-39. 2007
  2. pmc Xenobiotic metabolism, disposition, and regulation by receptors: from biochemical phenomenon to predictors of major toxicities
    Curtis J Omiecinski
    Department of Veterinary and Biomedical Sciences, Center for Molecular Toxicology and Carcinogenesis, Penn State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 120:S49-75. 2011
  3. pmc Bioassay-directed fractionation for discovery of bioactive neutral lipids guided by relative mass defect filtering and multiplexed collision-induced dissociation
    Michael C Stagliano
    Department of Chemistry, 104 Chemistry Building, The Pennsylvania State University, University Park, PA 16802, USA
    Rapid Commun Mass Spectrom 24:3578-84. 2010
  4. pmc Selective phthalate activation of naturally occurring human constitutive androstane receptor splice variants and the pregnane X receptor
    Joshua G DeKeyser
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 120:381-91. 2011
  5. pmc Differentiation of human embryonic stem cells along a hepatic lineage
    Stephanie M Zamule
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Chem Biol Interact 190:62-72. 2011
  6. pmc Chelation of lysosomal iron protects dopaminergic SH-SY5Y neuroblastoma cells from hydrogen peroxide toxicity by precluding autophagy and Akt dephosphorylation
    Roberta Castino
    Laboratorio di Patologia Molecolare, Centro di Biotecnologie per la Ricerca Medica Applicata, Dipartimento di Scienze Mediche, Universita del Piemonte Orientale A Avogadro, 28100 Novara, Italy
    Toxicol Sci 123:523-41. 2011
  7. pmc Multi-species analyses of direct activators of the constitutive androstane receptor
    Curtis J Omiecinski
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Penn State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 123:550-62. 2011
  8. pmc The orphan nuclear receptor DAX-1 functions as a potent corepressor of the constitutive androstane receptor (NR1I3)
    Elizabeth M Laurenzana
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA
    Mol Pharmacol 82:918-28. 2012
  9. pmc The human constitutive androstane receptor promotes the differentiation and maturation of hepatic-like cells
    Fengming Chen
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Dev Biol 384:155-65. 2013
  10. pmc Proteasomal interaction as a critical activity modulator of the human constitutive androstane receptor
    Tao Chen
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, 101 Life Sciences Building, Pennsylvania State University, University Park, PA 16802, U S A
    Biochem J 458:95-107. 2014

Detail Information

Publications22

  1. pmc CAR2 displays unique ligand binding and RXRalpha heterodimerization characteristics
    Scott S Auerbach
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Drug Metab Dispos 35:428-39. 2007
    ....
  2. pmc Xenobiotic metabolism, disposition, and regulation by receptors: from biochemical phenomenon to predictors of major toxicities
    Curtis J Omiecinski
    Department of Veterinary and Biomedical Sciences, Center for Molecular Toxicology and Carcinogenesis, Penn State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 120:S49-75. 2011
    ..This review will briefly summarize these developments and investigate the expanding roles of xenobiotic receptor biology in the underlying basis of toxicological response to chemical agents...
  3. pmc Bioassay-directed fractionation for discovery of bioactive neutral lipids guided by relative mass defect filtering and multiplexed collision-induced dissociation
    Michael C Stagliano
    Department of Chemistry, 104 Chemistry Building, The Pennsylvania State University, University Park, PA 16802, USA
    Rapid Commun Mass Spectrom 24:3578-84. 2010
    ..The identity of this compound was determined to be di(2-ethylhexyl) phthalate using GC/MS, and it was ranked as a promising candidate for reporter assay screening...
  4. pmc Selective phthalate activation of naturally occurring human constitutive androstane receptor splice variants and the pregnane X receptor
    Joshua G DeKeyser
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 120:381-91. 2011
    ..Alternative gene splicing results in variant CAR receptors that selectively recognize phthalates and BPA. The interaction of phthalates with CAR and PXR suggests a xenobiotic response that is complex and biologically redundant...
  5. pmc Differentiation of human embryonic stem cells along a hepatic lineage
    Stephanie M Zamule
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Chem Biol Interact 190:62-72. 2011
    ....
  6. pmc Chelation of lysosomal iron protects dopaminergic SH-SY5Y neuroblastoma cells from hydrogen peroxide toxicity by precluding autophagy and Akt dephosphorylation
    Roberta Castino
    Laboratorio di Patologia Molecolare, Centro di Biotecnologie per la Ricerca Medica Applicata, Dipartimento di Scienze Mediche, Universita del Piemonte Orientale A Avogadro, 28100 Novara, Italy
    Toxicol Sci 123:523-41. 2011
    ..The present work establishes HO(ยท) as the autophagy-inducing ROS and highlights the need for free lysosomal iron for its production within mitochondria in response to hydrogen peroxide...
  7. pmc Multi-species analyses of direct activators of the constitutive androstane receptor
    Curtis J Omiecinski
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Penn State University, University Park, Pennsylvania 16802, USA
    Toxicol Sci 123:550-62. 2011
    ....
  8. pmc The orphan nuclear receptor DAX-1 functions as a potent corepressor of the constitutive androstane receptor (NR1I3)
    Elizabeth M Laurenzana
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA
    Mol Pharmacol 82:918-28. 2012
    ..The results of this investigation identify DAX-1 as a novel and potent CAR corepressor and suggest that DAX-1 functions as a coordinate hepatic regulator of CAR's biological function...
  9. pmc The human constitutive androstane receptor promotes the differentiation and maturation of hepatic-like cells
    Fengming Chen
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Dev Biol 384:155-65. 2013
    ..Together, these results define a novel role for human CAR in hepatic lineage commitment. ..
  10. pmc Proteasomal interaction as a critical activity modulator of the human constitutive androstane receptor
    Tao Chen
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, 101 Life Sciences Building, Pennsylvania State University, University Park, PA 16802, U S A
    Biochem J 458:95-107. 2014
    ..Taken together, these data demonstrate that the proteasome complex functions at multiple levels to regulate the functional biology of hCAR activity. ..
  11. pmc Kynurenic acid is a potent endogenous aryl hydrocarbon receptor ligand that synergistically induces interleukin-6 in the presence of inflammatory signaling
    Brett C Dinatale
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16803, USA
    Toxicol Sci 115:89-97. 2010
    ..Our findings have established that KA is a potent AHR endogenous ligand that can induce IL6 production and xenobiotic metabolism in cells at physiologically relevant concentrations...
  12. pmc The uremic toxin 3-indoxyl sulfate is a potent endogenous agonist for the human aryl hydrocarbon receptor
    Jennifer C Schroeder
    Center for Molecular Toxicology and Carcinogenesis and Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, Pennsylvania 16802, USA
    Biochemistry 49:393-400. 2010
    ..Our results also suggest that prolonged activation of the AHR by I3S may contribute to toxicity observed in kidney dialysis patients and thus represent a possible therapeutic target...
  13. pmc Gene expression profiling of extracellular matrix as an effector of human hepatocyte phenotype in primary cell culture
    Jeanine L Page
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, 101 Life Sciences Building, The Pennsylvania State University, University Park, PA 16802, USA
    Toxicol Sci 97:384-97. 2007
    ....
  14. pmc Transactivation of a DR-1 PPRE by a human constitutive androstane receptor variant expressed from internal protein translation start sites
    Matthew A Stoner
    Center for Molecular Toxicology and Carcinogenesis, The Pennsylvania State University, University Park, PA 16802, USA
    Nucleic Acids Res 35:2177-90. 2007
    ....
  15. pmc Gene expression profiling and differentiation assessment in primary human hepatocyte cultures, established hepatoma cell lines, and human liver tissues
    Katy M Olsavsky
    Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Toxicol Appl Pharmacol 222:42-56. 2007
    ....
  16. pmc Aryl-hydrocarbon receptor activation regulates constitutive androstane receptor levels in murine and human liver
    Rushang D Patel
    Department of Veterinary and Biomedical Sciences and The Center for Molecular Toxicology and Carcinogenesis, The Pennsylvania State University, University Park, PA 16802, USA
    Hepatology 46:209-18. 2007
    ..Finally, CAR was also up-regulated in primary human hepatocytes in response to AhR activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin and benzo[a]pyrene...
  17. pmc Regulation of the human cathepsin E gene by the constitutive androstane receptor
    Jeanine L Page
    Center for Molecular Toxicology and Carcinogenesis and the Department of Veterinary and Biomedical Sciences, 101 Life Sciences Building, The Pennsylvania State University, University Park, PA 16802, USA
    Arch Biochem Biophys 467:132-8. 2007
    ..Thus, these studies demonstrate CAR-mediated regulation of CTSE within primary hepatocyte cultures from several individual donors and suggest that elevated CTSE activity may play a functional role in the etiology of hepatocarcinogenesis...
  18. pmc Preservation of hepatic phenotype in lentiviral-transduced primary human hepatocytes
    Stephanie M Zamule
    Center for Molecular Toxicology and Carcinogenesis and Department of Veterinary and Biomedical Sciences, Pennsylvania State University, 101 Life Sciences Building, University Park, PA 16802, USA
    Chem Biol Interact 173:179-86. 2008
    ....
  19. pmc Expression profiling of interindividual variability following xenobiotic exposures in primary human hepatocyte cultures
    Katy M O Goyak
    Department of Veterinary and Biomedical Sciences, Penn State University, University Park, PA 16802, USA
    Toxicol Appl Pharmacol 231:216-24. 2008
    ....
  20. pmc Di(2-ethylhexyl) phthalate is a highly potent agonist for the human constitutive androstane receptor splice variant CAR2
    Joshua G DeKeyser
    Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, The Pennsylvania State University, University Park, PA 16802, USA
    Mol Pharmacol 75:1005-13. 2009
    ..The discovery that CAR2 is an ultimate human DEHP receptor identifies a novel pathway modulating human DEHP toxicity with potential clinical implications for a subset of patients undergoing critical care medical interventions...
  21. pmc Effect of rifampin and nelfinavir on the metabolism of methadone and buprenorphine in primary cultures of human hepatocytes
    David E Moody
    Center for Human Toxicology, Department of Pharmacology and Toxicology, University of Utah, Salt Lake City, UT 84108, USA
    Drug Metab Dispos 37:2323-9. 2009
    ....
  22. pmc Phenobarbital and propiconazole toxicogenomic profiles in mice show major similarities consistent with the key role that constitutive androstane receptor (CAR) activation plays in their mode of action
    Richard A Currie
    Syngenta Ltd, Jealott s Hill International Research Centre, Bracknell, Berkshire RG42 6EY, UK Electronic address
    Toxicology 321:80-8. 2014
    ....