PPARα Induces the Expression of CAR That Works as a Negative Regulator of PPARα Functions in Mouse Livers.

The nuclear receptor peroxisome proliferator-activated receptor α (PPARα) is a transcription factor that controls the transcription of genes responsible for fatty acid metabolism. We have recently reported a possible drug-drug interaction mechanism via the interaction of PPARα with the xenobiotic nuclear receptor constitutive androstane receptor (CAR). Drug-activated CAR competes with the transcriptional coactivator against PPARα and prevents PPARα-mediated lipid metabolism. In this study, to elucidate the crosstalk between CAR and PPARα, we focused on the influence of PPARα activation on CAR's gene expression and activation. Male C57BL/6N mice (8-12 weeks old, n = 4) were treated with PPARα and CAR activators (fenofibrate and phenobarbital, respectively), and hepatic mRNA levels were determined using quantitative reverse transcription PCR. Reporter assays using the mouse Car promoter were performed in HepG2 cells to determine the PPARα-dependent induction of CAR. CAR KO mice were treated with fenofibrate, and the hepatic mRNA levels of PPARα target genes were determined. Treatment of mice with a PPARα activator increased Car mRNA levels as well as genes related to fatty acid metabolism. In reporter assays, PPARα induced the promoter activity of the Car gene. Mutation of the putative PPARα-binding motif prevented PPARα-dependent induction of reporter activity. In electrophoresis mobility shift assay, PPARα bound to the DR1 motif of the Car promoter. Since CAR has been reported to attenuate PPARα-dependent transcription, CAR was considered a negative feedback protein for PPARα activation. Treatment with fenofibrate induced the mRNA levels of PPARα target genes in Car-null mice more than those in wild-type mice, suggesting that CAR functions as a negative feedback factor for PPARα.

PXR Suppresses PPARα-Dependent HMGCS2 Gene Transcription by Inhibiting the Interaction between PPARα and PGC1α.

BACKGROUND: PXR is a xenobiotic-responsive nuclear receptor that controls the expression of drug-metabolizing enzymes. Drug-induced activation of PXR sometimes causes drug-drug interactions due to the induced metabolism of co-administered drugs. Our group recently reported a possible drug-drug interaction mechanism via an interaction between the nuclear receptors CAR and PPARα. As CAR and PXR are structurally and functionally related receptors, we investigated possible crosstalk between PXR and PPARα. METHODS: Human hepatocyte-like HepaRG cells were treated with various PXR ligands, and mRNA levels were determined by quantitative reverse transcription PCR. Reporter assays using the HMGCS2 promoter containing a PPARα-binding motif and mammalian two-hybrid assays were performed in HepG2 or COS-1 cells. RESULTS: Treatment with PXR activators reduced the mRNA levels of PPARα target genes in HepaRG cells. In reporter assays, PXR suppressed PPARα-dependent gene expression in HepG2 cells. In COS-1 cells, co-expression of PGC1α, a common coactivator of PPARα and PXR, enhanced PPARα-dependent gene transcription, which was clearly suppressed by PXR. Consistently, in mammalian two-hybrid assays, the interaction between PGC1α and PPARα was attenuated by ligand-activated PXR. CONCLUSION: The present results suggest that ligand-activated PXR suppresses PPARα-dependent gene expression by inhibiting PGC1α recruitment.

Antiepileptic Drug-Activated Constitutive Androstane Receptor Inhibits Peroxisome Proliferator-Activated Receptor α and Peroxisome Proliferator-Activated Receptor γ Coactivator 1α-Dependent Gene Expression to Increase Blood Triglyceride Levels.

Long-term administration of some antiepileptic drugs often increases blood lipid levels. In this study, we investigated its molecular mechanism by focusing on the nuclear receptors constitutive active/androstane receptor (CAR) and peroxisome proliferator-activated receptor α (PPARα), which are key transcription factors for enzyme induction and lipid metabolism, respectively, in the liver. Treatment of mice with the CAR activator phenobarbital, an antiepileptic drug, increased plasma triglyceride levels and decreased the hepatic expression of PPARα target genes related to lipid metabolism. The increase in PPARα target gene expression induced by fenofibrate, a PPARα ligand, was inhibited by cotreatment with phenobarbital. CAR suppressed PPARα-dependent gene transcription in HepG2 cells but not in COS-1 cells. The mRNA level of peroxisome proliferator-activated receptor γ coactivator 1α (PGC1α), a coactivator for both CAR and PPARα, in COS-1 cells was much lower than in HepG2 cells. In reporter assays with COS-1 cells overexpressing PGC1α, CAR suppressed PPARα-dependent gene transcription, depending on the coactivator-binding motif. In mammalian two-hybrid assays, CAR attenuated the interaction between PGC1α and PPARα Chemical inhibition of PGC1α prevented phenobarbital-dependent increases in plasma triglyceride levels and the inhibition of PPARα target gene expression. These results suggest that CAR inhibits the interaction between PPARα and PGC1α, attenuating PPARα-dependent lipid metabolism. This might explain the antiepileptic drug-induced elevation of blood triglyceride levels. SIGNIFICANCE STATEMENT: Constitutive active/androstane receptor activated by antiepileptic drugs inhibits the peroxisome proliferator-activated receptor α-dependent transcription of genes related to lipid metabolism and upregulates blood triglyceride levels. The molecular mechanism of this inhibition involves competition between these nuclear receptors for coactivator peroxisome proliferator-activated receptor γ coactivator-1α binding.