Genetic Determinants of Junin Virus Attenuation

Abstract

The New World arenavirus Junin (JUNV) is the causative agent of the Argentine hemorrhagic fever (AHF), a deadly disease endemic to central regions of Argentina. The live-attenuated Candid #1 strain of JUNV is currently used to vaccinate human population at risk. However, the mechanism of attenuation of this strain is still largely unknown. Therefore, the identification and functional characterization of viral genetic factors implicated in JUNV pathogenesis or attenuation would significantly improve the understanding of the molecular mechanisms underlying AHF and facilitate the development of novel, effective and safe vaccines. To this end, an RNA polymerase I/II-based reverse genetics system was utilized to rescue the wild type pathogenic Romero and attenuated Candid #1 strains of JUNV. Both recombinant viruses exhibited similar in vitro growth kinetics and in vivo biological properties to their parental counterparts. This system was further used to generate chimeric JUNV variants encoding different gene combinations of Romero and Candid #1. Analysis of virulence of the chimeric viruses in a guinea pigs model of lethal infection that closely reproduces the features of AHF, identified the envelope glycoproteins (GPs) as the major determinants of pathogenesis and attenuation of JUNV. Therefore, the chimeric viruses expressing the GPs of Romero and Candid #1 exhibited virulent and attenuated phenotypes in guinea pigs, respectively. Comparison of the transcriptional and protein expression profiles of the chimeric JUNV variants demonstrated marked differences in the levels of viral RNA synthesis and protein expression between the attenuated and virulent JUNV variants. Further analysis showed that the GPC of Candid #1 undergoes abnormal post-translational modification and induces endoplasmic reticulum (ER) stress, which may facilitate immune recognition of JUNV infection. In addition, the small RING finger protein Z that is a negative regulator of the viral polymerase complex was retained in infected cells when it was coexpressed with the GPs of Romero, but not the GPs of Candid #1, therefore suggesting a molecular mechanism for the higher levels of viral RNA synthesis in cells infected with attenuated JUNV variants expressing the GPC of Candid #1. Thus, these findings provided further insights into the mechanism of JUNV attenuation.