Author(s): TESHOME MEBATSION, MATTHIAS J SCHNELL, KARLKLAUS CONZELMANN
A reverse genetics approach which allows the generation of infectious defective rabies virus (RV) particles entirely from plasmid-encoded genomes and proteins (K.K.Conzelmann and M.Schnell, J.Virol.68:713–719, 1994) was used to investigate the ability of a heterologous lyssa virus glycoprotein (G) and chimeric G constructs to function in the formation of infectious RV-like particles. Virions containing a chloramphenicol acetyltransferase (CAT) reporter gene (SDI-CAT) were generated in cells simultaneously expressing the genomic RNA analog, the RVN, P, M, and L proteins, and engineered G constructs from transfected plasmids. The infectivity of particles was determined by a CAT assay after passage to helper virus-infected cells. The heterologous G protein from Eth-16 virus (Mokola virus, lyssa virus serotype 3) as well as a construct in which the ecto domain of RV G was fused to the cytoplasmic and transmembrane domains of the Eth-16 virus G rescued infectious SDI-CAT particles. In contrast, a chimeric protein composed of the amino-terminal half of the Eth-16 virus G and the carboxy-terminal half of RV G failed to produce infectious particles. Site directed mutagenesis was used to convert the antigenics ite III of RVG to the corresponding sequence of Eth-16G. This chimeric protein rescued infectious SDI-CAT particles as efficiently as RV G. Virions containing the chimeric protein were specifically neutralized by an anti-Eth-16 virus serum and escaped neutralization by a monoclonal antibody directed against RV antigenic siteIII. The results show that entire structural domains as well as short surface epitopes of lyssa virus G proteins may be exchanged without affecting the structure required to mediate infection of cells.