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Journal of Virology, August 2003, p. 8924-8933, Vol. 77, No. 16
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.16.8924-8933.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Incorporation of Tick-Borne Encephalitis Virus Replicons into Virus-Like Particles by a Packaging Cell Line

Rainer Gehrke, Michael Ecker,{dagger} Stephan W. Aberle, Steven L. Allison, Franz X. Heinz, and Christian W. Mandl*

Institute of Virology, University of Vienna, Vienna, Austria

Received 6 February 2003/ Accepted 19 May 2003

RNA replicons derived from flavivirus genomes show considerable potential as gene transfer and immunization vectors. A convenient and efficient encapsidation system is an important prerequisite for the practical application of such vectors. In this work, tick-borne encephalitis (TBE) virus replicons and an appropriate packaging cell line were constructed and characterized. A stable CHO cell line constitutively expressing the two surface proteins prM/M and E (named CHO-ME cells) was generated and shown to efficiently export mature recombinant subviral particles (RSPs). When replicon Nd{Delta}ME lacking the prM/M and E genes was introduced into CHO-ME cells, virus-like particles (VLPs) capable of initiating a single round of infection were released, yielding titers of up to 5 x 107/ml in the supernatant of these cells. Another replicon (Nd{Delta}CME) lacking the region encoding most of the capsid protein C in addition to proteins prM/M and E was not packaged by CHO-ME cells. As observed with other flavivirus replicons, both TBE virus replicons appeared to exert no cytopathic effect on their host cells. Sedimentation analysis revealed that the Nd{Delta}ME-containing VLPs were physically distinct from RSPs and similar to infectious virions. VLPs could be repeatedly passaged in CHO-ME cells but maintained the property of being able to initiate only a single round of infection in other cells during these passages. CHO-ME cells can thus be used both as a source for mature TBE virus RSPs and as a safe and convenient replicon packaging cell line, providing the TBE virus surface proteins prM/M and E in trans.

* Corresponding author. Mailing address: Institute of Virology, Kinderspitalgasse 15, A-1095 Vienna, Austria. Phone: 43 1 404 90, ext. 79502. Fax: 43 1 404 90 9795. E-mail: christian.mandl{at}univie.ac.at.

{dagger} Present address: Baxter AG, A-1220 Vienna, Austria.

Journal of Virology, August 2003, p. 8924-8933, Vol. 77, No. 16
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.16.8924-8933.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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