Influenza NA and PB1 Gene Segments Interact during the Formation of Viral Progeny: Localization of the Binding Region within the PB1 Gene.

The influenza A virus genome comprises eight negative-sense viral RNAs (vRNAs) that form individual ribonucleoprotein (RNP) complexes. In order to incorporate a complete set of each of these vRNAs, the virus uses a selective packaging mechanism that facilitates co-packaging of specific gene segments but whose molecular basis is still not fully understood. Recently, we used a competitive transfection model where plasmids encoding the A/Puerto Rico/8/34 (PR8) and A/Udorn/307/72 (Udorn) PB1 gene segments were competed to show that the Udorn PB1 gene segment is preferentially co-packaged into progeny virions with the Udorn NA gene segment. Here we created chimeric PB1 genes combining both Udorn and PR8 PB1 sequences to further define the location within the Udorn PB1 gene that drives co-segregation of these genes and show that nucleotides 1776-2070 of the PB1 gene are crucial for preferential selection. In vitro assays examining specific interactions between Udorn NA vRNA and purified vRNAs transcribed from chimeric PB1 genes also supported the importance of this region in the PB1-NA interaction. Hence, this work identifies an association between viral genes that are co-selected during packaging. It also reveals a region potentially important in the RNP-RNP interactions within the supramolecular complex that is predicted to form prior to budding to allow one of each segment to be packaged in the viral progeny. Our study lays the foundation to understand the co-selection of specific genes, which may be critical to the emergence of new viruses with pandemic potential.

Selective packaging of the influenza A genome and consequences for genetic reassortment.

Influenza A viruses package their segmented RNA genome in a selective manner. Electron tomography, biochemical assays, and replication assays of viruses produced by reverse genetics recently unveiled molecular details of this mechanism, whereby different influenza viral strains form different and unique networks of direct intermolecular RNA-RNA interactions. Together with detailed views of the three-dimensional structure of the viral ribonucleoparticles, these recent advances help us understand the rules that govern genome packaging. They also have deep implications for the genetic reassortment processes, which are responsible for devastating pandemics.

Empaquetage sélectif du génome des influenzavirus A.

Electron microscopy of influenza A virus (IAV) and three-dimensional reconstruction of their interior by electron tomography, combined with genetic, biochemical and virology assays, has revealed that genome packaging of IAVs is a selective process, the molecular mechanisms of which start to be unveiled. The eight genomic viral RNAs (vRNAs) most likely form a supramolecular complex maintained by base-pairings within the strain-dependent packaging signals of each vRNA. Visualization of viral ribonucleoproteins inside cells also brought new insights about spatio-temporal assembly of the supramolecular complexes, prior to their incorporation into budding virions. Altogether, these data improve our understanding of the rules governing packaging of the IAV genome and offer clues for optimization of vaccine seeds production. Genetic reassortment events between different IAVs, which can lead to severe pandemics, are probably also affected by the rules that govern genome packaging.