Constitutive Androstane Receptor-1 (CAR-1; NR1I3i1)
Human Constitutive Androstane Receptor-1 (CAR-1) exhibits constitutive activity, but can be further regulated through direct ligand interactions. The primary application of this reporter assay system is in the screening of test samples to quantify any functional inverse-agonist activity that they may exert on human CAR-1. INDIGO’s CAR-1 Reporter Assay System utilizes proprietary mammalian cells engineered to provide high-level expression of human CAR-1 (NR1I3 isoform 1), which is a constitutively active transcription factor.
Reporter Cells utilize a modified version of human CAR1 in which the N-terminal DNA binding domain (DBD) has been replaced with the GAL4-DBD. The human CAR1 ligand binding domain (LBD) is unaltered and fully functional. These reporter cells also incorporate a luciferase cDNA functionally linked to the GAL4-upstream activation sequence (UAS). Thus, quantifying expressed luciferase activity provides a sensitive surrogate measure of changes in CAR1 activity resulting from a direct interaction between a treatment compound and the nuclear receptor. Because this assay system expresses a GAL4-DBD + hCAR1 LBD hybrid receptor, the bio-activity of activators that act through indirect mechanisms (such as those that alter the phosphorylation status of the native N-terminal amino acid sequence of CAR1) may be dampened or go undetected.
CAR is encoded by the NR1I3 gene. This gene encodes a member of the nuclear receptor superfamily, and is a key regulator of xenobiotic and endobiotic metabolism. The protein binds to DNA as a monomer or a heterodimer with the retinoid X receptor and regulates the transcription of target genes involved in drug metabolism and bilirubin clearance, such as cytochrome P450 family members. Unlike most nuclear receptors, this transcriptional regulator is constitutively active in the absence of ligand but is regulated by both agonists and inverse agonists. Ligand binding results in translocation of this protein to the nucleus, where it activates or represses target gene transcription. These ligands include bilirubin, a variety of foreign compounds, steroid hormones, and prescription drugs.
For more information on CAR, visit the Nuclear Receptor Resource.
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CAR-1 Reporter Cells are prepared using INDIGO’s proprietary CryoMite™ process. This cryo-preservation method yields high cell viability post-thaw, and provides the convenience of immediately dispensing healthy, division-competent reporter cells into assay plates. There is no need for intermediate spin-and-wash steps, viability determinations, or cell titer adjustments.
Kits are offered in different assay formats to accommodate researchers' needs: 3x 32 and 1x 96 assay formats for screening small numbers of test compounds, as well as custom bulk reagents for HTS applications. Assay systems are all inclusive, providing reporter cells, optimized growth media, media for diluting test compounds, a positive-control agonist, luciferase detection reagent, a white assay plate, a detailed protocol, and a protocol quick guide. All kits are shipped on dry ice.
The principle application of this assay product is in the screening of test samples to quantify functional inverse-agonist activities that they may exert against the human constitutive androstane receptor. This kit product is an all-inclusive assay system that includes, in addition to CAR-1 Reporter Cells, two optimized media for use during cell culture and (optionally) in diluting the test samples, a positive-control inverse-agonist, Luciferase Detection Reagent, a cell culture-ready assay plate, and a detailed protocol.
| Product Family | Product Number | Product Description |
| IB0091 CAR-1 (NR1I3) | IB00911-32 | Human CAR-1 Reporter Assay System, 3 x 32 assays in 96-well format |
| IB00911 | Human CAR-1 Reporter Assay System, 1 x 96-well format assays |
Service Assays: Human, Rat, Mouse
The primary application of INDIGO’s cell-based nuclear receptor assays are to quantitatively assess the bioactivity of a test compound as an agonist (activator) or antagonist (inhibition of an agonist response) of a given receptor. Service assays include a positive control reference compound and ‘vehicle’ control for every experiment. A formal study report and all data files are provided to the client upon completion of the study. To receive a quote for your proposed study, complete & submit the online “Request a Quote” form or contact an INDIGO Customer Service Representative to discuss your desired study parameters. To initiate a Service Study, download and complete all fields of the Excel worksheet “Service Work Order" then submit the electronic file to INDIGO Customer Service.
Sedaxane—Use of Nuclear Receptor Transactivation Assays, Toxicogenomics, and Toxicokinetics as Part of a Mode of Action Framework for Rodent Liver Tumors
ABSTRACT Experimental data demonstrate a mode of action (MOA) for liver tumors in male rats and mice treated with sedaxane that starts with activation of CAR, followed by altered expression of CAR-responsive genes, increased cell proliferation, and eventually clonal expansion of preneoplastic cells, leading to the development of altered foci and tumors. This MOA isRead More
Profiling Drug Activity of Human and Ortholog Xenobiotic-Sensing Receptors: PXR, CAR, AhR, and FXR
View Full Size Research conducted by: Koji Toyokawa (1), Ewa Maddox (1), Jack Vanden Huevel (1,2), & Bruce Sherf (1) (1) INDIGO Biosciences, Inc., 1981 Pine Hall Rd, State College, PA, USA (2) Center for Molecular Toxicology and Carcinogenesis, 325 Life Sciences Building, Penn State University, University Park, PA 16802, USA Date of Publication: 2017
Differential modulation of FXR activity by chlorophacinone and ivermectin analogs
ABSTRACT Chemicals that alter normal function of farnesoid X receptor (FXR) have been shown to affect the homeostasis of bile acids, glucose, and lipids. Several structural classes of environmental chemicals and drugs that modulated FXR transactivation were previously identified by quantitative high-throughput screening (qHTS) of the Tox21 10 K chemical collection. In the present study, weRead More
Mouse Liver Tumor Mode of Action: Use of Toxicogenomics in Weight of Evidence for Human Relevance Assessment
ABSTRACT The Use of Toxicogenomics in Chemical Safety Testing Current safety testing is geared to produce and accept descriptive data from high-dose animal studies Interpretation of this information has effectively protected our health and safety for decates; however, hindered by lack of understanding mechanistic information Concern over chemical safety for humans and the environment hasRead More
Effects of Munitions Compounds on Xenobiotic-Activated Nuclear Receptors and Signaling Pathways
ABSTRACT Exposure to certain munitions compounds is know to alter physiological functions in test organisms, however little is known about their molecular and cellular effects. Several nuclear receptors are regulated by xenobiotic compounds. These nuclear receptors belong to a class of ligand-activated transcription factors that, when heterodimerized with RXRa and bound to their respective DNARead More
Indigo Announces In Vitro Toxicology Platform
In vitro toxicology platform provides predictive model of liver toxicity. Aims to reduce the high rates of drug-induced liver damage State College, PA (May 4, 2016) INDIGO Biosciences, the recognized industry leader in nuclear receptor research, has completed development of an in vitro toxicology platform, meeting the demand for predictive liver toxicity models. INDIGO’s inRead More
Oxysterols are agonist ligands of RORγt and drive Th17 cell differentiation
ABSTRACT The RAR-related orphan receptor gamma t (RORγt) is a nuclear receptor required for generating IL-17–producing CD4+ Th17 T cells, which are essential in host defense and may play key pathogenic roles in autoimmune diseases. Oxysterols elicit profound effects on immune and inflammatory responses as well as on cholesterol and lipid metabolism. Here, we describe the identification of several naturally occurringRead More
Evidence for triclosan-induced activation of human and rodent xenobiotic nuclear receptors
ABSTRACT The bacteriostat triclosan (2,4,4′-trichloro-2′-hydroxydiphenylether) (TCS) decreases rat serum thyroxine via putative nuclear receptor (NR) interaction(s) and subsequent transcriptional up-regulation of hepatic catabolism and clearance. However, due to the evolutionary divergence of the constitutive androstane and pregnane-X receptors (CAR, PXR), TCS-mediated downstream effects may be species-dependent. To test the hypothesis that TCS activates xenobiotic NRsRead More