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Journal of Virology, June 2002, p. 5974-5984, Vol. 76, No. 12
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.12.5974-5984.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Expression of Hepatitis C Virus Proteins Induces Distinct Membrane Alterations Including a Candidate Viral Replication Complex

Denise Egger,1 Benno Wölk,2 Rainer Gosert,1,2 Leonardo Bianchi,3 Hubert E. Blum,2 Darius Moradpour,2 and Kurt Bienz1*

Institute for Medical Microbiology, University of Basel, CH-4003 Basel, and Emeritus,1 University of Basel, CH-4114 Hofstetten, Switzerland,3 Department of Medicine II, University of Freiburg, D-79106 Freiburg, Germany2

Received 14 December 2001/ Accepted 22 March 2002

Plus-strand RNA viruses characteristically replicate their genome in association with altered cellular membranes. In the present study, the capacity of hepatitis C virus (HCV) proteins to elicit intracellular membrane alterations was investigated by expressing, in tetracycline-regulated cell lines, a comprehensive panel of HCV proteins individually as well as in the context of the entire HCV polyprotein. As visualized by electron microscopy (EM), expression of the combined structural proteins core-E1-E2-p7, the NS3-4A complex, and protein NS4B induced distinct membrane alterations. By immunogold EM (IEM), the membrane-altering proteins were always found to localize to the respective altered membranes. NS4B, a protein of hitherto unknown function, induced a tight structure, designated membranous web, consisting of vesicles in a membranous matrix. Expression of the entire HCV polyprotein gave rise to membrane budding into rough endoplasmic reticulum vacuoles, to the membranous web, and to tightly associated vesicles often surrounding the membranous web. By IEM, all HCV proteins were found to be associated with the NS4B-induced membranous web, forming a membrane-associated multiprotein complex. A similar web-like structure in livers of HCV-infected chimpanzees was previously described (Pfeifer et al., Virchows Arch. B., 33:233-243, 1980). In view of this finding and the observation that all HCV proteins accumulate on the membranous web, we propose that the membranous web forms the viral replication complex in HCV-infected cells.

* Corresponding author. Mailing address: Institute for Medical Microbiology, University of Basel, Petersplatz 10, CH-4003 Basel, Switzerland. Phone: 41 61 267 3290. Fax: 41 61 267 3283. E-mail: Kurt.Bienz{at}unibas.ch.

Journal of Virology, June 2002, p. 5974-5984, Vol. 76, No. 12
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.12.5974-5984.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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