title: Incorporation of tick-borne encephalitis virus replicons into virus-like particles by a packaging cell line.
abstract: 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 NdDeltaME 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 10(7)/ml in the supernatant of these cells. Another replicon (NdDeltaCME) 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 NdDeltaME-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.
authors.affiliation: Institute of Virology, University of Vienna, Vienna, Austria.
authors.forename: Rainer
authors.initials: R
authors.lastname: Gehrke
authors.forename: Michael
authors.initials: M
authors.lastname: Ecker
authors.forename: Stephan W
authors.initials: SW
authors.lastname: Aberle
authors.forename: Steven L
authors.initials: SL
authors.lastname: Allison
authors.forename: Franz X
authors.initials: FX
authors.lastname: Heinz
authors.forename: Christian W
authors.initials: CW
authors.lastname: Mandl
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D017931:DNA Primers; D004669:Encephalitis Viruses, Tick-Borne / Q000235:genetics / Q000502:physiology*; D012093:Replicon; D019065:Virus Assembly
chemical_list: D017931:DNA Primers