Autor(es): Rozen-Gagnon Kathryn, Stapleford Kenneth A, Mongelli Vanesa, Blanc Hervé, Failloux Anna-Bella, Saleh Maria-Carla, Vignuzzi Marco

Resumo: Arboviruses cycle through both vertebrates - invertebrates, which requires them to adapt to disparate hosts while maintaining genetic integrity during genome replication. To study the genetic mechanisms - determinants of these processes, we use chikungunya virus (CHIKV), a re-emerging human pathogen transmitted by the Aedes mosquito. We previously isolated a high fidelity (or antimutator) polymerase variant, C483Y, which had decreased fitness in both mammalian - mosquito hosts, suggesting this residue may be a key molecular determinant. To further investigate effects of position 483 on RNA-dependent RNA-polymerase (RdRp) fidelity, we substituted every amino acid at this position. We isolated novel mutators with decreased replication fidelity - higher mutation frequencies, allowing us to examine the fitness of error-prone arbovirus variants. Although CHIKV mutators displayed no major replication defects in mammalian cell culture, they had reduced specific infectivity - were attenuated in vivo. Unexpectedly, mutator phenotypes were suppressed in mosquito cells - the variants exhibited significant defects in RNA synthesis. Consequently, these replication defects resulted in strong selection for reversion during infection of mosquitoes. Since residue 483 is conserved among alphaviruses, we examined the analogous mutations in Sindbis virus (SINV), which also reduced polymerase fidelity - generated replication defects in mosquito cells. However, replication defects were mosquito cell-specific - were not observed in Drosophila S2 cells, allowing us to evaluate the potential attenuation of mutators in insect models where pressure for reversion was absent. Indeed, the SINV mutator variant was attenuated in fruit flies. These findings confirm that residue 483 is a determinant regulating alphavirus polymerase fidelity - demonstrate proof of principle that arboviruses can be attenuated in mammalian - insect hosts by reducing fidelity.

Imprenta: PLoS Pathogens, v. 10, n. 1, p. e1003877, 2014

Identificador do objeto digital: 10.1371/journal.ppat.1003877

Descritores: Chikungunya virus - Arbovirus ; Chikungunya virus - Biosynthesis ; Chikungunya virus - Cell ; Chikungunya virus - Genome ; Chikungunya virus - Pathogenesis ; Chikungunya virus - Proteins ; Chikungunya virus - RNA ; Chikungunya Virus - Virus

Data de publicação: 2014