title: Transcriptional regulation of human ferredoxin reductase through an intronic enhancer in steroidogenic cells.
abstract: Ferredoxin reductase (FDXR, also known as adrenodoxin reductase) is a mitochondrial flavoprotein that transfers electrons from NADPH to mitochondrial cytochrome P450 enzymes, mediating the function of an iron-sulfur cluster protein, ferredoxin. FDXR functions in various metabolic processes including steroidogenesis. It is well known that multiple steroidogenic enzymes are regulated by a transcription factor steroidogenic factor-1 (SF-1, also known as Ad4BP). Previously, we have shown that SF-1 transduction causes human mesenchymal stem cell differentiation into steroidogenic cells. Genome-wide analysis of differentiated cells, using a combination of DNA microarray and promoter tiling array analyses, showed that FDXR is a novel SF-1 target gene. In this study, the transcriptional regulatory mechanism of FDXR was examined in steroidogenic cells. A chromatin immunoprecipitation assay revealed that a novel SF-1 binding region was located within intron 2 of the human FDXR gene. Luciferase reporter assays showed that FDXR transcription was activated through the novel SF-1 binding site within intron 2. Endogenous SF-1 knockdown in human adrenocortical H295R and KGN cells decreased FDXR expression. In H295R cells, strong binding of two histone markers of active enhancers, histones H3K27ac and H3K4me2, were detected near the SF-1 binding site within intron 2. Furthermore, the binding of these histone markers was decreased concurrent with SF-1 knockdown in H295R cells. These results indicated that abundant FDXR expression in these steroidogenic cells was maintained through SF-1 binding to the intronic enhancer of the FDXR gene.
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan; Translational Research Center, Organization for Life Science Advancement Programs, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Yoshitaka
authors: Y
authors: Imamichi
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan; Translational Research Center, Organization for Life Science Advancement Programs, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan. Electronic address: mizutani@u-fukui.ac.jp.
authors: Tetsuya
authors: T
authors: Mizutani
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Yunfeng
authors: Y
authors: Ju
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Takehiro
authors: T
authors: Matsumura
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan; Translational Research Center, Organization for Life Science Advancement Programs, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Shinya
authors: S
authors: Kawabe
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Masafumi
authors: M
authors: Kanno
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan; Translational Research Center, Organization for Life Science Advancement Programs, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Takashi
authors: T
authors: Yazawa
authors: Department of Biochemistry, Faculty of Medical Sciences, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan; Translational Research Center, Organization for Life Science Advancement Programs, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Eiheiji, Fukui 910-1193, Japan.
authors: Kaoru
authors: K
authors: Miyamoto
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D004268:DNA-Binding Proteins; D004742:Enhancer Elements, Genetic; D005287:Ferredoxin-NADP Reductase / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D006657:Histones / Q000235:genetics; D006801:Humans; D007438:Introns; D056484:Jumonji Domain-Containing Histone Demethylases / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D054339:Steroidogenic Factor 1 / Q000235:genetics* / Q000378:metabolism; D013256:Steroids / Q000096:biosynthesis / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D004268:DNA-Binding Proteins; D006657:Histones; D054339:Steroidogenic Factor 1; D013256:Steroids; D056484:Jumonji Domain-Containing Histone Demethylases; C524220:KDM6B protein, human; D005287:Ferredoxin-NADP Reductase
keywords: ChIP; EMSA; FDX1; FDX2; FDXR; Ferredoxin reductase; GAPDH; Histone modification; MSCs; NR5A1; P450s; RT-PCR; SF-1; STAR; Steroidogenic cell; Steroidogenic factor-1; TSS; Transcription factor; Transcriptional regulation; chromatin immunoprecipitation; cytochrome P450 enzymes; electrophoresis mobility shift assay; ferredoxin 1; ferredoxin 2; ferredoxin reductase; glyceraldehyde-3-phosphate dehydrogenase; mesenchymal stem cells; nuclear receptor subfamily 5 group A member 1; reverse transcriptase-polymerase chain reaction; steroidogenic acute regulatory protein; steroidogenic factor-1; transcription start site