Orphan Nuclear Receptors Constitutive Androstane Receptor and Pregnane X Receptor Share Xenobiotic and Steroid Ligands

doi:10.1074/jbc.M001215200

Orphan Nuclear Receptors Constitutive Androstane Receptor and Pregnane X Receptor Share Xenobiotic and Steroid Ligands*

From the Departments of ^‡Molecular Endocrinology,^§Molecular Biochemistry, ^¶Molecular Sciences, and ^**Medicinal Chemistry Glaxo Wellcome Research and Development, Research Triangle Park, North Carolina 27709 and the ^‖Department of Clinical Pharmacology and Storr Liver Unit, University of Sydney at Westmead Hospital, Westmead, New South Wales 2145, Australia

Abstract

Xenobiotics induce the transcription of cytochromes P450 (CYPs) 2B and 3A through the constitutive androstane receptor (CAR; NR1I3) and pregnane X receptor (PXR; NR1I2), respectively. In this report, we have systematically compared a series of xenobiotics and natural steroids for their effects on mouse and human CAR and PXR. Our results demonstrate dual regulation of PXR and CAR by a subset of compounds that affect CYP expression. Moreover, there are marked pharmacological differences between the mouse (m) and human (h) orthologs of both CAR and PXR. For example, the planar hydrocarbon 1,4-bis[2-(3,5-dichloropyridyl-oxy)]benzene activates mCAR and hPXR but has little or no activity on hCAR and mPXR. In contrast, the CAR deactivator androstanol activates both mouse and human PXR. Similarly, the PXR activator clotrimazole is a potent deactivator of hCAR. Using radioligand binding and fluorescence resonance energy transfer assays, we demonstrate that several of the compounds that regulate mouse and human CAR, including natural steroids, bind directly to the receptors. Our results suggest that CAR, like PXR, is a steroid receptor that is capable of recognizing structurally diverse compounds. Moreover, our findings underscore the complexity in the physiologic response to xenobiotics.

Footnotes

↵* The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‡ To whom correspondence should be addressed: Glaxo Wellcome Research and Development, 5 Moore Dr., Research Triangle Park, NC 27709. Tel.: 919-483-5601; Fax: 919-483-6147; E-mail: sak15922@glaxowellcome.com.
Published, JBC Papers in Press, March 20, 2000, DOI 10.1074/jbc.M001215200
↵2 B. Goodwin and C. Liddle, unpublished data.
Abbreviations:

CYP

cytochrome P450

TCPOBOP

1,4-bis[2-(3,5-dichloropyridyloxy)]benzene

CAR

constitutive androstane receptor

PXR

pregnane X receptor

FRET

fluorescence resonance energy transfer

LBD

ligand binding domain

DTT

dithiothreitol

PB

phenobarbital

RXR

9-cis-retinoic acid receptor

TBS

Tris-buffered saline

SPA

scintillation proximity binding assay

CHAPS

3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid

PCN

pregnenolone 16α-carbonitrile
- Received February 14, 2000.
- Revision received March 17, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.