title: Modulation of Dcytb (Cybrd 1) expression and function by iron, dehydroascorbate and Hif-2α in cultured cells.
abstract: BACKGROUND
  Duodenal cytochrome b (Dcytb) is a mammalian plasma ferric reductase enzyme that catalyses the reduction of ferric to ferrous ion in the process of iron absorption. The current study investigates the relationship between Dcytb, iron, dehydroascorbate (DHA) and Hif-2α in cultured cell lines.
  
  
  METHODS
  Dcytb and Hif-2α protein expression was analysed by Western blot technique while gene regulation was determined by quantitative PCR. Functional analyses were carried out by ferric reductase and (59)Fe uptake assays.
  
  
  RESULTS
  Iron and dehydroascorbic acid treatment of cells inhibited Dcytb mRNA and protein expression. Desferrioxamine also enhanced Dcytb mRNA level after cells were treated overnight. Dcytb knockdown in HuTu cells resulted in reduced mRNA expression and lowered reductase activity. Preloading cells with DHA (to enhance intracellular ascorbate levels) did not stimulate reductase activity fully in Dcytb-silenced cells, implying a Dcytb-dependence of ascorbate-mediated ferrireduction. Moreover, Hif-2α knockdown in HuTu cells led to a reduction in reductase activity and iron uptake.
  
  
  CONCLUSIONS
  Taken together, this study shows the functional regulation of Dcytb reductase activity by DHA and Hif-2α.
  
  
  CONCLUSIONS
  Dcytb is a plasma membrane protein that accepts electrons intracellularly from DHA/ascorbic acid for ferrireduction at the apical surface of cultured cells and enterocytes.
authors: Diabetes and Nutritional Sciences Division, King's College London, London, UK.
authors: Xiaomin
authors: X
authors: Luo
authors: Melanie
authors: M
authors: Hill
authors: Anna
authors: A
authors: Johnson
authors: Gladys O
authors: GO
authors: Latunde-Dada
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D003573:Cytochrome b Group / Q000235:genetics / Q000378:metabolism*; D003683:Dehydroascorbic Acid / Q000494:pharmacology*; D004379:Duodenal Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D038181:FMN Reductase / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D007501:Iron / Q000494:pharmacology*; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D003573:Cytochrome b Group; D003683:Dehydroascorbic Acid; D038181:FMN Reductase; D007501:Iron; D010088:Oxidoreductases; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000099162:Endothelial PAS Domain-Containing Protein 1; C422926:CYBRD1 protein, human; C023903:ferric citrate iron reductase
keywords: AA; ARNT; Aryl hydrocarbon receptor nuclear translocator; Ascorbic acid; Cells; DFO; DHA; Dcytb; Dehydroascorbic acid; Desferrioxamine; Duodenal cytochrome b; FAC; Fe-NTA; Ferric ammonium citrate; Ferric-nitrilotriacetate; Gene silencing; HRE; Hif-2α; Hypoxia response element; PHDs; Prolyl hydroxylases; VHL; Von–Hippel–Lindau