Volker Kinast1, Agnieszka Plociennikowska2, Thilo Bracht3, Daniel Todt1, Richard J P Brown4, Tujana Boldanova5, Yudi Zhang6, Yannick Brüggemann7, Martina Friesland6, Michael Engelmann1, Gabrielle Vieyres6, Ruth Broering8, Florian W R Vondran9, Markus H Heim5, Barbara Sitek3, Ralf Bartenschlager10, Thomas Pietschmann11, Eike Steinmann12. 1. Institute of Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany; Faculty of Medicine, Department for Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany. 2. Department of Infectious Diseases, Molecular Virology, Heidelberg University, Heidelberg, Germany; Division Virus-Associated Carcinogenesis, German Cancer Research Center, Heidelberg, Germany. 3. Medizinisches Proteom-Center, Ruhr University Bochum, Bochum, Germany. 4. Institute of Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany; Division of Veterinary Medicine, Paul Ehrlich Institute, Langen, Germany. 5. Department of Biomedicine, University of Basel and Division of Gastroenterology and Hepatology, University Hospital Basel, Basel, Switzerland. 6. Institute of Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany. 7. Faculty of Medicine, Department for Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany. 8. Department of Gastroenterology and Hepatology, University Hospital Essen, University Duisburg-Essen, Essen, Germany. 9. ReMediES, Department of General, Visceral and Transplantation Surgery, Hannover Medical School, Hannover, Germany; German Centre for Infection Research (DZIF), partner site Hannover-Braunschweig, Hannover, Germany. 10. Department of Infectious Diseases, Molecular Virology, Heidelberg University, Heidelberg, Germany; Division Virus-Associated Carcinogenesis, German Cancer Research Center, Heidelberg, Germany; German Center for Infection Research (DZIF), partner site Heidelberg, Heidelberg, Germany. 11. Institute of Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany; German Centre for Infection Research (DZIF), partner site Hannover-Braunschweig, Hannover, Germany. 12. Institute of Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany; Faculty of Medicine, Department for Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany. Electronic address: eike.steinmann@ruhr-uni-bochum.de.
Abstract
BACKGROUND & AIMS: HCV is a positive-strand RNA virus that primarily infects human hepatocytes. Recent studies have reported that C19orf66 is expressed as an interferon (IFN)-stimulated gene; however, the intrinsic regulation of this gene within the liver as well as its antiviral effects against HCV remain elusive. METHODS: Expression of C19orf66 was quantified in both liver biopsies and primary human hepatocytes, with or without HCV infection. Mechanistic studies of the potent anti-HCV phenotype mediated by C19orf66 were conducted using state-of-the-art virological, biochemical and genetic approaches, as well as correlative light and electron microscopy and transcriptome and proteome analysis. RESULTS: Upregulation of C19orf66 mRNA was observed in both primary human hepatocytes upon HCV infection and in the livers of patients with chronic hepatitis C (CHC). In addition, pegIFNα/ribavirin therapy induced C19orf66 expression in patients with CHC. Transcriptomic profiling and whole cell proteomics of hepatoma cells ectopically expressing C19orf66 revealed no induction of other antiviral genes. Expression of C19orf66 restricted HCV infection, whereas CRIPSPR/Cas9 mediated knockout of C19orf66 attenuated IFN-mediated suppression of HCV replication. Co-immunoprecipitation followed by mass spectrometry identified a stress granule protein-dominated interactome of C19orf66. Studies with subgenomic HCV replicons and an expression system revealed that C19orf66 expression impairs HCV-induced elevation of phosphatidylinositol-4-phosphate, alters the morphology of the viral replication organelle (termed the membranous web) and thereby targets viral RNA replication. CONCLUSION: C19orf66 is an IFN-stimulated gene, which is upregulated in hepatocytes within the first hours post IFN treatment or HCV infection in vivo. The encoded protein possesses specific antiviral activity against HCV and targets the formation of the membranous web. Our study identifies C19orf66 as an IFN-inducible restriction factor with a novel antiviral mechanism that specifically targets HCV replication. LAY SUMMARY: Interferon-stimulated genes are thought to be important to for antiviral immune responses to HCV. Herein, we analysed C19orf66, an interferon-stimulated gene, which appears to inhibit HCV replication. It prevents the HCV-induced elevation of phosphatidylinositol-4-phosphate and alters the morphology of HCV's replication organelle.
BACKGROUND & AIMS:HCV is a positive-strand RNA virus that primarily infects human hepatocytes. Recent studies have reported that C19orf66 is expressed as an interferon (IFN)-stimulated gene; however, the intrinsic regulation of this gene within the liver as well as its antiviral effects against HCV remain elusive. METHODS: Expression of C19orf66 was quantified in both liver biopsies and primary human hepatocytes, with or without HCV infection. Mechanistic studies of the potent anti-HCV phenotype mediated by C19orf66 were conducted using state-of-the-art virological, biochemical and genetic approaches, as well as correlative light and electron microscopy and transcriptome and proteome analysis. RESULTS: Upregulation of C19orf66 mRNA was observed in both primary human hepatocytes upon HCV infection and in the livers of patients with chronic hepatitis C (CHC). In addition, pegIFNα/ribavirin therapy induced C19orf66 expression in patients with CHC. Transcriptomic profiling and whole cell proteomics of hepatoma cells ectopically expressing C19orf66 revealed no induction of other antiviral genes. Expression of C19orf66 restricted HCV infection, whereas CRIPSPR/Cas9 mediated knockout of C19orf66 attenuated IFN-mediated suppression of HCV replication. Co-immunoprecipitation followed by mass spectrometry identified a stress granule protein-dominated interactome of C19orf66. Studies with subgenomic HCV replicons and an expression system revealed that C19orf66 expression impairs HCV-induced elevation of phosphatidylinositol-4-phosphate, alters the morphology of the viral replication organelle (termed the membranous web) and thereby targets viral RNA replication. CONCLUSION:C19orf66 is an IFN-stimulated gene, which is upregulated in hepatocytes within the first hours post IFN treatment or HCV infection in vivo. The encoded protein possesses specific antiviral activity against HCV and targets the formation of the membranous web. Our study identifies C19orf66 as an IFN-inducible restriction factor with a novel antiviral mechanism that specifically targets HCV replication. LAY SUMMARY: Interferon-stimulated genes are thought to be important to for antiviral immune responses to HCV. Herein, we analysed C19orf66, an interferon-stimulated gene, which appears to inhibit HCV replication. It prevents the HCV-induced elevation of phosphatidylinositol-4-phosphate and alters the morphology of HCV's replication organelle.
Authors: Natasha W Hanners; Katrina B Mar; Ian N Boys; Jennifer L Eitson; Pamela C De La Cruz-Rivera; R Blake Richardson; Wenchun Fan; Mary Wight-Carter; John W Schoggins Journal: Proc Natl Acad Sci U S A Date: 2021-12-07 Impact factor: 12.779