title: Different mechanisms of hepatitis C virus RNA polymerase activation by cyclophilin A and B in vitro.
abstract: BACKGROUND
  Cyclophilins (CyPs) are cellular proteins that are essential to hepatitis C virus (HCV) replication. Since cyclosporine A was discovered to inhibit HCV infection, the CyP pathway contributing to HCV replication is a potential attractive stratagem for controlling HCV infection. Among them, CyPA is accepted to interact with HCV nonstructural protein (NS) 5A, although interaction of CyPB and NS5B, an RNA-dependent RNA polymerase (RdRp), was proposed first.
  
  
  METHODS
  CyPA, CyPB, and HCV RdRp were expressed in bacteria and purified using combination column chromatography. HCV RdRp activity was analyzed in vitro with purified CyPA and CyPB.
  
  
  RESULTS
  CyPA at a high concentration (50× higher than that of RdRp) but not at low concentration activated HCV RdRp. CyPB had an allosteric effect on genotype 1b RdRp activation. CyPB showed genotype specificity and activated genotype 1b and J6CF (2a) RdRps but not genotype 1a or JFH1 (2a) RdRps. CyPA activated RdRps of genotypes 1a, 1b, and 2a. CyPB may also support HCV genotype 1b replication within the infected cells, although its knockdown effect on HCV 1b replicon activity was controversial in earlier reports.
  
  
  CONCLUSIONS
  CyPA activated HCV RdRp at the early stages of transcription, including template RNA binding. CyPB also activated genotype 1b RdRp. However, their activation mechanisms are different.
  
  
  CONCLUSIONS
  These data suggest that both CyPA and CyPB are excellent targets for the treatment of HCV 1b, which shows the greatest resistance to interferon and ribavirin combination therapy.
authors.affiliation: Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, People's Republic of China.
authors.forename: Leiyun
authors.initials: L
authors.lastname: Weng
authors.forename: Xiao
authors.initials: X
authors.lastname: Tian
authors.forename: Yayi
authors.initials: Y
authors.lastname: Gao
authors.forename: Koichi
authors.initials: K
authors.lastname: Watashi
authors.forename: Kunitada
authors.initials: K
authors.lastname: Shimotohno
authors.forename: Takaji
authors.initials: T
authors.lastname: Wakita
authors.forename: Michinori
authors.initials: M
authors.lastname: Kohara
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Toyoda
mesh_terms: D021984:Cyclophilin A / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D021983:Cyclophilins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic; D005838:Genotype; D016174:Hepacivirus / Q000201:enzymology / Q000235:genetics*; D006526:Hepatitis C / Q000378:metabolism* / Q000821:virology; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D066298:In Vitro Techniques; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic; D014779:Virus Replication
chemical_list: D007166:Immunosuppressive Agents; D050505:Mutant Proteins; D012367:RNA, Viral; C081786:cyclophilin B; D016572:Cyclosporine; D012324:RNA-Dependent RNA Polymerase; D021984:Cyclophilin A; D021983:Cyclophilins