Resource allocation during Flavivirus lifecycle

Abstract

Single-stranded RNA virus infection cycle progresses by resource (viral RNA) allocation to segregated macromolecular complexes: host ribosomes (translation), viral RNA polymerase (replication) and viral structural proteins (assembly). Many positive-strand (+) RNA viruses from Flaviviridae family further compartmentalize the replication process in membranous vesicles, which helps in selective resource allocation to viral RNA polymerase. We hypothesized that the spatial segregation of replication process played a key role in regulating viral RNA lifecycle, as suggested by its conservation across a large number of (+)RNA viruses.

Using Japanese encephalitis virus (JEV) infection in Neuro2a cells, we measured dynamics of resource distribution during viral life cycle and observed high levels of viral RNA translation (60%) during early stages of infection. Virus infection of the cell then progresses to a phase where replication and virus packaging dominate in spite of minimal perturbations in global translation. We further evaluate translation dynamics on JEV genome using ribosome profiling to understand the resource allocation strategies. The assay also allowed us to assess elongation rate of ribosomes and indicates a possible role of upstream ORF expression along with tRNA levels in potentially regulating translation efficiency. To integrate our findings of virus lifecyle dynamics, we next employ a deterministic, process-based cell infection model in which replication is spatially segregated from translation and replication vesicles grow logistically, to best explain our experimental data. Together, our complementary experimental and modeling approaches predicts replication compartmentalization as a critical event for a successful intracellular flaviviral infection.