1887

Abstract

A -packaging system for hepatitis C virus (HCV) replicons lacking envelope glycoproteins was developed. The replicons were efficiently encapsidated into infectious particles after expression of homologous HCV envelope proteins under the control of an adenoviral vector. Interestingly, expression of core or core, p7 and NS2 with envelope proteins did not enhance encapsidation. Expression of heterologous envelope proteins, in the presence or absence of heterologous core, p7 and NS2, did not rescue single-round infectious particle production. To increase the titre of homologous, single-round infectious particles in our system, successive cycles of encapsidation and infection were performed. Four cycles resulted in a 100-fold increase in the yield of particles. Sequence analysis revealed a total of 16 potential adaptive mutations in two independent experiments. Except for a core mutation in one experiment, all the mutations were located in non-structural regions mainly in NS5A (four in domain III and two near the junction with the NS5B gene). Reverse genetics studies suggested that D2437A and S2443T adaptive mutations, which are located at the NS5A-B cleavage site did not affect viral replication, but enhanced the single-round infectious particles assembly only -encapsidation model. In conclusion, our encapsidation system enables the production of HCV single-round infectious particles. This system is adaptable and can positively select variants. The adapted variants promote encapsidation and should constitute a valuable tool in the development of replicon-based HCV vaccines.

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2013-05-01
2019-10-16
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