title: Ca(2+) /S100 proteins regulate HCV virus NS5A-FKBP8/FKBP38 interaction and HCV virus RNA replication.
abstract: OBJECTIVE
  FKBP8/FKBP38 is a unique FK506-binding protein with a C-terminal membrane anchor and localizes at the outer membranes of mitochondria and the endoplasmic reticulum. Similar to some immunophilins, such as FKBP51, FKBP52 and Cyclophilin 40, FKBP8/FKBP38 contain a putative Calmodulin-binding domain and a tetratricopeptide-repeat (TPR) domain for the binding of Hsp90. Both Hsp90 and the non-structural protein 5A (NS5A) of the hepatitis C virus (HCV) interact specifically with FKBP8/FKBP38 through its TPR domain, and the ternary complex formation plays a critical role in HCV RNA replication. The goal of this study is to evaluate that the host factor inhibits the ternary complex formation and the replication of HCV in vitro and in vivo.
  
  
  METHODS
  S100 proteins, FKBP38, FKBP8, HCV NS5A, Hsp90, and calmodulin were expressed in E.coli and purified. In vitro binding studies were performed by GST pull-down, S-tag pull-down and surface plasmon resonance analyses. The effect of S100 proteins on HCV replication was analysed by Western blotting using an HCV NS3 antibody following transfection of S100 proteins into the HCV replicon harbouring cell line (sO cells).
  
  
  RESULTS
  In vitro binding studies showed that S100A1, S100A2, S100A6, S100B and S100P directly interacted with FKBP8/FKBP38 in a Ca(2+) -dependent manner and inhibited the FKBP8/FKBP38-Hsp90 and FKBP8/FKBP38-NS5A interactions. Furthermore, overexpression of S100A1, S100A2 and S100A6 in sO cells resulted in the efficient inhibition of HCV replication.
  
  
  CONCLUSIONS
  The association of the S100 proteins with FKBP8/FKBP38 provides a novel Ca(2+) -dependent regulatory role in HCV replication through the NS5A-host protein interaction.
authors.affiliation: Department of Gastroenterology and Neurology, Kagawa University Faculty of Medicine, Kagawa, Japan.
authors.forename: Joji
authors.initials: J
authors.lastname: Tani
authors.forename: Seiko
authors.initials: S
authors.lastname: Shimamoto
authors.forename: Kyoko
authors.initials: K
authors.lastname: Mori
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Kato
authors.forename: Kohji
authors.initials: K
authors.lastname: Moriishi
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Matsuura
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Tokumitsu
authors.forename: Mitsumasa
authors.initials: M
authors.lastname: Tsuchiya
authors.forename: Tomohito
authors.initials: T
authors.lastname: Fujimoto
authors.forename: Kiyohito
authors.initials: K
authors.lastname: Kato
authors.forename: Hisaaki
authors.initials: H
authors.lastname: Miyoshi
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Masaki
authors.forename: Ryoji
authors.initials: R
authors.lastname: Kobayashi
mesh_terms: D002118:Calcium / Q000378:metabolism*; D004926:Escherichia coli; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D016174:Hepacivirus / Q000235:genetics / Q000502:physiology*; D006801:Humans; D010957:Plasmids / Q000235:genetics; D012367:RNA, Viral / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000378:metabolism; D009418:S100 Proteins / Q000378:metabolism*; D020349:Surface Plasmon Resonance; D022021:Tacrolimus Binding Proteins / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase
chemical_list: D002118:Calcium; D018841:HSP90 Heat-Shock Proteins; D012367:RNA, Viral; D011994:Recombinant Proteins; D009418:S100 Proteins; D022021:Tacrolimus Binding Proteins; D017361:Viral Nonstructural Proteins; C471205:FKBP8 protein, human; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase
keywords: Ca2+-binding proteins; HCV replicon harbouring cells; Hsp90; ternary complex formation; tetratricopeptide repeat