title: Utilization of RNA polymerase I promoter and terminator sequences to develop a DNA transfection system for the study of hepatitis C virus internal ribosomal entry site-dependent translation.
abstract: BACKGROUND
  Hepatitis C virus (HCV) causes severe liver diseases in a large population worldwide. HCV protein translation is controlled by an internal ribosomal entry site (IRES) within the 5'-untranslated region (UTR). HCV IRES-dependent translation is critical for HCV-associated pathogenesis.
  
  
  OBJECTIVE
  To develop a plasmid DNA transfection system by using RNA polymerase I promoter and terminator sequences for studying HCV IRES-dependent translation.
  
  
  METHODS
  A gene cassette containing HCV 5'-UTR, Renilla luciferase reporter gene, and HCV 3'-UTR was inserted between RNA polymerase I promoter and terminator sequences. HCV IRES-directed translation was determined by luciferase assay after transfection.
  
  
  RESULTS
  Transfection of the RNA polymerase I-HCV IRES plasmid into human hepatoma Huh-7 and HepG2 cells resulted in luciferase gene expression. Deletion of the IIIf domain in HCV IRES dramatically reduced luciferase activity.
  
  
  CONCLUSIONS
  Our results indicated that the plasmid vector system-based on RNA polymerase I promoter and terminator sequences represents an effective approach for the study of HCV IRES-dependent translation.
authors.affiliation: Vaccine and Infectious Disease Organization (VIDO), University of Saskatchewan, 120 Veterinary Road, Saskatoon, Sask., Canada S7N 5E3.
authors.forename: Jae-Ku
authors.initials: JK
authors.lastname: Oem
authors.forename: Zhonghua
authors.initials: Z
authors.lastname: Xiang
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhou
authors.forename: Lorne A
authors.initials: LA
authors.lastname: Babiuk
authors.forename: Qiang
authors.initials: Q
authors.lastname: Liu
mesh_terms: D020121:5' Untranslated Regions; D016174:Hepacivirus / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008156:Luciferases; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012318:RNA Polymerase I / Q000235:genetics* / Q000378:metabolism; D012367:RNA, Viral; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*; D013728:Terminator Regions, Genetic; D014162:Transfection
chemical_list: D020121:5' Untranslated Regions; D012367:RNA, Viral; D008156:Luciferases; D012318:RNA Polymerase I