title: Identification of the IFITM3 gene as an inhibitor of hepatitis C viral translation in a stable STAT1 cell line.
abstract: To investigate the functions of signal transducers and activators of transcription 1 (STAT1)-induced anti-hepatitis C viral (HCV) effects, a stable Huh7.5 cell line (Huh7.5-STAT1ER) was established that constitutively expresses a fusion protein (STAT1ER) of STAT1 and the mouse oestrogen receptor (ER), which forms STAT1ER homodimers after 4-hydroxytamoxifen (4-HT) treatment. This inducible and cytokine/receptor-independent STAT1 activation system allowed us to investigate the anti-HCV effects of STAT1ER activation after inducing IFN-stimulated gene (ISG) expression. The anti-HCV effects of dimerized STAT1ER fusion protein were determined by real-time PCR in a time-dependent fashion post-HCV (JFH-1) infection. HCV (JFH-1) RNA decreased 48% at 72 h after 4-HT treatment. To distinguish the inhibitory effects of STAT1ER activation on HCV RNA replication or HCV internal ribosomal entry site (IRES)-mediated translation, a dicistronic pRL-HL construct was used in the studies. Both cellular (Cap-dependent) and HCV IRES-mediated (Cap-independent) translation were decreased by 63% and 57% at 72 h post-STAT1ER activation in the STAT1ER cell line. In our previous studies, interferon-induced transmembrane protein 3 [(IFITM3) (1-8U)] was found to inhibit HCV RNA replication. Subsequently, elevated expression of the 1-8U gene was confirmed by Western blotting in the Huh7.5-STAT1ER cell line. To further investigate the 1-8U function with both in vivo and in vitro studies, the 1-8U gene was found to suppress cellular and HCV IRES-mediated translation.
authors.affiliation: Division of Gastroenterology, Department of Medicine, College of Medicine-Jacksonville, University of Florida, Jacksonville, FL, USA.
authors.forename: L
authors.initials: L
authors.lastname: Yao
authors.forename: H
authors.initials: H
authors.lastname: Dong
authors.forename: H
authors.initials: H
authors.lastname: Zhu
authors.forename: D
authors.initials: D
authors.lastname: Nelson
authors.forename: C
authors.initials: C
authors.lastname: Liu
authors.forename: L
authors.initials: L
authors.lastname: Lambiase
authors.forename: X
authors.initials: X
authors.lastname: Li
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D015870:Gene Expression; D016174:Hepacivirus / Q000276:immunology*; D006801:Humans; D008565:Membrane Proteins / Q000276:immunology*; D051379:Mice; D014176:Protein Biosynthesis; D055503:Protein Multimerization; D012367:RNA, Viral / Q000096:biosynthesis; D016601:RNA-Binding Proteins / Q000276:immunology*; D060888:Real-Time Polymerase Chain Reaction; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D013629:Tamoxifen / Q000031:analogs & derivatives / Q000378:metabolism; D015533:Transcriptional Activation; D014764:Viral Proteins / Q000096:biosynthesis*
chemical_list: D002352:Carrier Proteins; C080082:IFITM3 protein, human; D008565:Membrane Proteins; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D011960:Receptors, Estrogen; D011993:Recombinant Fusion Proteins; D050794:STAT1 Transcription Factor; D014764:Viral Proteins; C010140:estrophilin; D013629:Tamoxifen; C016601:afimoxifene