Abstract
Nucleoprotein (N) is a key element in rabies virus (RABV) replication. To further investigate the effect of N on RABV, we manipulated an infectious cDNA clone of the RABV HEP-Flury to rearrange the N gene from its wild-type position of 1 (N-P-M-G-L) to 2 (P-N-M-G-L), 3 (P-M-N-G-L), or 4 (P-M-G-N-L), using an approach that left the viral nucleotide sequence unaltered. Subsequently, viable viruses were recovered from each of the rearranged cDNA and examined for their gene expression levels, growth kinetics in cell culture, pathogenicity in suckling mice and protection in mice. The results showed that gene rearrangement decreased N mRNA transcription and vRNA replication. As a result, all viruses with rearranged genomes showed worse replication than that of rHEP-Flury in NA cells at a MOI of 0.01, but equivalent or slightly better replication levels at a MOI of 3. Consequently, the lethality in suckling mice infected with N4 was clearly attenuated compared with rHEP-Flury. However, the protection to mice was not enhanced. This study not only gives us insight into the understanding of the phenotype of RABV N gene rearrangement, but also helps with rabies vaccine candidate construction.
Highlights
Gene rearrangement has been used to alter the genotypes of viruses that result in predictable changes in gene expression, which can be invaluable for gene function and control studies [1]
Previous work has demonstrated that rearranging the five viral genes of vesicular stomatitis virus (VSV), a prototype Vesiculovirus genus of the Rhabdoviridae family, could affect its relative protein expression levels and alter phenotypes and lethality in mice infected with recombinant viruses [1,2]
Gene of HEP-Flury was studied by rearranging the N gene positions from proto-site (1st site) to 2nd, The recovered were designated as N1 aspects, and N4 according the specific
Summary
Gene rearrangement has been used to alter the genotypes of viruses that result in predictable changes in gene expression, which can be invaluable for gene function and control studies [1]. Previous work has demonstrated that rearranging the five viral genes of vesicular stomatitis virus (VSV), a prototype Vesiculovirus genus of the Rhabdoviridae family, could affect its relative protein expression levels and alter phenotypes and lethality in mice infected with recombinant viruses [1,2]. Neither of the RNA type in rearranged viruses infected cells, nor the relative molar ratio of the proteins in mature virus particles were changed by gene rearrangement [3]. The Rabies virus (RABV), which is a member of the genus Lyssavirus of the family Rhabdoviridae, causes severe neurological disease and death in almost all kinds of mammals, including humans [4]. The RABV N protein, which is decisive for virus replication, tightly encapsidates the genomic and anti-genomic RNA of RABV to form the viral
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