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Replication-Competent Recombinant Vesicular Stomatitis Virus Encoding Hepatitis C Virus Envelope Proteins

Hideki Tani,¹ Yasumasa Komoda,^1,2 Eiko Matsuo,^1, Kensuke Suzuki,^1,2 Itsuki Hamamoto,^1, Tetsuo Yamashita,¹ Kohji Moriishi,¹ Kazuhito Fujiyama,³ Tatsuya Kanto,⁴ Norio Hayashi,⁴ Ania Owsianka,⁵ Arvind H. Patel,⁵ Michael A. Whitt,⁶ and Yoshiharu Matsuura^1*

Department of Molecular Virology, Research Institute for Microbial Diseases,¹ The International Center for Biotechnology,³ Department of Gastroenterology and Hepatology, Graduate School of Medicine, Osaka University, Osaka, Japan,⁴ Central Pharmaceutical Research Institute, Japan Tobacco Inc., Osaka, Japan,² MRC Virology Unit, Institute of Virology, Glasgow, United Kingdom,⁵ Department of Molecular Sciences, University of Tennessee Health Science Center, Memphis, Tennessee⁶

Received 22 March 2007/ Accepted 21 May 2007

Although in vitro replication of the hepatitis C virus (HCV) JFH1 clone of genotype 2a (HCVcc) has been developed, a robust cell culture system for the 1a and 1b genotypes, which are the most prevalent viruses in the world and resistant to interferon therapy, has not yet been established. As a surrogate virus system, pseudotype viruses transiently bearing HCV envelope proteins based on the vesicular stomatitis virus (VSV) and retrovirus have been developed. Here, we have developed a replication-competent recombinant VSV with a genome encoding unmodified HCV E1 and E2 proteins in place of the VSV envelope protein (HCVrv) in human cell lines. HCVrv and a pseudotype VSV bearing the unmodified HCV envelope proteins (HCVpv) generated in 293T or Huh7 cells exhibited high infectivity in Huh7 cells. Generation of infectious HCVrv was limited in some cell lines examined. Furthermore, HCVrv but not HCVpv was able to propagate and form foci in Huh7 cells. The infection of Huh7 cells with HCVpv and HCVrv was neutralized by anti-hCD81 and anti-E2 antibodies and by sera from chronic HCV patients. The infectivity of HCVrv was inhibited by an endoplasmic reticulum -glucosidase inhibitor, N-(n-nonyl) deoxynojirimycin (Nn-DNJ), but not by a Golgi mannosidase inhibitor, deoxymannojirimycin. Focus formation of HCVrv in Huh7 cells was impaired by Nn-DNJ treatment. These results indicate that the HCVrv developed in this study can be used to study HCV envelope proteins with respect to not only the biological functions in the entry process but also their maturation step.

Published ahead of print on 6 June 2007.

Present address: London School of Hygiene and Tropical Medicine, University of London, London, United Kingdom.

Present address: Infectious Disease Surveillance Center, National Institute of Infectious Diseases, Tokyo, Japan.

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