Abstract
Within influenza virus infected cells, viral genomic RNA are selectively packed into progeny virions, which predominantly contain a single copy of 8 viral RNA segments. Intersegmental RNA-RNA interactions are thought to mediate selective packaging of each viral ribonucleoprotein complex (vRNP). Clear evidence of a specific interaction network culminating in the full genomic set has yet to be identified. Using multi-color fluorescence in situ hybridization to visualize four vRNP segments within a single cell, we developed image-based models of vRNP-vRNP spatial dependence. These models were used to construct likely sequences of vRNP associations resulting in the full genomic set. Our results support the notion that selective packaging occurs during cytoplasmic transport and identifies the formation of multiple distinct vRNP sub-complexes that likely form as intermediate steps toward full genomic inclusion into a progeny virion. The methods employed demonstrate a statistically driven, model based approach applicable to other interaction and assembly problems.
Highlights
Influenza A virus, part of the orthomyxoviridae family, has a segmented genome of eight distinct viral RNA segments coding at least 11 major proteins and several auxiliary peptides
Influenza virus consists of eight viral ribonucleoproteins that are assembled by infected cells to produce new virions
We used images from a previous study in which up to four viral ribonucleoprotein complex (vRNP) had been visualized in the same cell to construct spatial point process models that measure how well the subcellular distribution of one vRNP can be predicted from one or more other vRNPs
Summary
Influenza A virus, part of the orthomyxoviridae family, has a segmented genome of eight distinct viral RNA segments coding at least 11 major proteins and several auxiliary peptides. Most notable of the 11 viral proteins are hemagglutinin (HA) and neurominidase (NA), the canonical H and N in influenza strain designation. A segmented genome offers potential evolutionary advantages during viral replication, in the form of reassortment. The exchange of genomic material between two distinct viral strains in a co-infected cell often causes a shift within the genome, greatly increasing viral genetic diversity. Segmented genomes come with an inherent mechanistic challenge: ensuring that progeny receive a complete set of genomic segments. Evidence suggests that there exists an active mechanism of selective packaging ensuring progeny viability through efficient and accurate genomic packaging [3, 4]
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