title: Transcription of the human microsomal epoxide hydrolase gene (EPHX1) is regulated by an HNF-4α/CAR/RXR/PSF complex.
abstract: Microsomal epoxide hydrolase (mEH) is a bifunctional protein that plays a central role in the metabolism of numerous xenobiotics as well as mediating the sodium-dependent transport of bile acids into hepatocytes where they are involved in cholesterol excretion and metabolism, lipid digestion and regulating numerous signaling pathways. Previous studies have demonstrated the critical role of GATA-4 and a C/EBPα-NF/Y complex in the regulation of the mEH gene (EPHX1). In this study we show that HNF-4α and CAR/RXR also bind to the proximal promoter region and regulate EPHX1 expression. Bile acids, which inhibit the expression of HNF-4α also decrease the expression of EPHX1. Studies also established that the binding of HNF-4α was essential for the activation of EPHX1 activity by CAR suggesting the formation of a complex between these adjacent factors. The nature of this regulatory complex was further explored using a biotinylated oligonucleotide of this region in conjunction with BioMag beads and mass spectrometric analysis which demonstrated the presence of an additional inhibitory factor (PSF), confirmed by co-immunoprecipitation and ChIP analyses, which interacted with DNA-bound CAR/RXR/HNF-4α forming a 4-component regulatory complex.
authors.affiliation: University of Southern California, Keck School of Medicine, Department of Biochemistry and Molecular Biology, 2011 Zonal Avenue, Los Angeles, CA 90033, USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Peng
authors.forename: Qin-Shi
authors.initials: QS
authors.lastname: Zhu
authors.forename: Shuping
authors.initials: S
authors.lastname: Zhong
authors.forename: Daniel
authors.initials: D
authors.lastname: Levy
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D002846:Chromatography, Affinity; D000090702:Constitutive Androstane Receptor; D024202:Electrophoretic Mobility Shift Assay; D004851:Epoxide Hydrolases / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D006367:HeLa Cells; D056945:Hep G2 Cells; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D000072670:PTB-Associated Splicing Factor; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D047488:Retinoid X Receptors / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000090702:Constitutive Androstane Receptor; C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; D000072670:PTB-Associated Splicing Factor; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D047488:Retinoid X Receptors; D004851:Epoxide Hydrolases
keywords: CAR; CDCA; ChIP; EMSA; EPHX1; HNF-4α; PSF; RXR; SHP; Transcriptional regulation; chenodeoxycholic acid; chromatin immunoprecipitation; constitutive androstane receptor; electrophoretic mobility shift assay; hepatocyte nuclear factor-4α; mEH; microsomal epoxide hydrolase; microsomal epoxide hydrolase gene; polypyrimidine tract binding protein-associated splicing factor; retinoid X receptor; small heterodimer partner