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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., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Influenza A are nuclear replicating viruses which hijack host machineries in order to achieve optimal infection. Numerous functional virus-host interactions have now been characterized, but little information has been gathered concerning their link to the virally induced remodeling of the host cellular architecture. In this study, we infected cells with several human and avian influenza viruses and we have analyzed their ultrastructural modifications by using electron and confocal microscopy. We discovered that infections lead to a major and systematic disruption of nucleoli and the formation of a large number of diverse viral structures showing specificity that depended on the subtype origin and genomic composition of viruses. We identified NS1 and M1 proteins as the main actors in the remodeling of the host ultra-structure and our results suggest that each influenza A virus strain could be associated with a specific cellular fingerprint, possibly correlated to the functional properties of their viral components., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Influenza is one of the most common infectious diseases in humans occurring as seasonal epidemic and sporadic pandemic outbreaks. The ongoing infections of humans with avian H5N1 influenza A viruses (IAV) and the past 2009 pandemic caused by the quadruple human/avian/swine reassortant (H1N1) virus highlights the permanent threat caused by these viruses. This review aims to describe the interaction between the virus and the host, with a particular focus on the role of proteases and HLA-G in the pathogenicity of influenza viruses., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Classical antiviral therapies target viral proteins and are consequently subject to resistance. To counteract this limitation, alternative strategies have been developed that target cellular factors. We hypothesized that such an approach could also be useful to identify broad-spectrum antivirals. The influenza A virus was used as a model for its viral diversity and because of the need to develop therapies against unpredictable viruses as recently underlined by the H1N1 pandemic. We proposed to identify a gene-expression signature associated with infection by different influenza A virus subtypes which would allow the identification of potential antiviral drugs with a broad anti-influenza spectrum of activity. We analyzed the cellular gene expression response to infection with five different human and avian influenza A virus strains and identified 300 genes as differentially expressed between infected and non-infected samples. The most 20 dysregulated genes were used to screen the connectivity map, a database of drug-associated gene expression profiles. Candidate antivirals were then identified by their inverse correlation to the query signature. We hypothesized that such molecules would induce an unfavorable cellular environment for influenza virus replication. Eight potential antivirals including ribavirin were identified and their effects were tested in vitro on five influenza A strains. Six of the molecules inhibited influenza viral growth. The new pandemic H1N1 virus, which was not used to define the gene expression signature of infection, was inhibited by five out of the eight identified molecules, demonstrating that this strategy could contribute to identifying new broad anti-influenza agents acting on cellular gene expression. The identified infection signature genes, the expression of which are modified upon infection, could encode cellular proteins involved in the viral life cycle. This is the first study showing that gene expression-based screening can be used to identify antivirals. Such an approach could accelerate drug discovery and be extended to other pathogens.

We have previously shown a clear differential genetic evolution of the hemagglutinin (HA) of human AH1N1 and AH1N2 viruses, isolated in southern France between 2001 and 2004. However, our analysis revealed that one single AH1N2 isolate, detected in 2003 (A/Lyon/0838/2003), had its HA clustering within the HAs of the AH1N1 subtypes. To determine if this virus was a new reassortant, the nucleotide sequences of its eight RNA gene segments were compared with those of five representative strains of the AH1N1, AH1N2 and AH3N2 viruses, isolated during the same time-period. According to the sequences obtained, the neuraminidase (NA) gene segment of the A/Lyon/0838/2003 (H1N2) virus was genetically closely related to those of the AH1N2 viruses, whereas the six internal genes appeared to be clustering with those of the AH1N1 viruses. This suggested that the A/Lyon/0838/2003 (H1N2) virus was the result of a second reassortment event that had occurred during the winter of 2002-2003; the N2 gene segment of an AH1N2 virus being introduced into an AH1N1 genetic background. Subsequently, we analysed the extremities of each gene segment of the viruses from the different subtypes, and experimentally reproduced, in vitro, AH1N2 reassortant viruses through co-infection of Madin-Darby canine kidney (MDCK) cells with both AH1N1 and AH1N2 viruses, isolated in the 2002-2003 influenza season. While the comparison of the AH1N1 and the AH1N2 gene segment extremities revealed no major differences, we successfully reproduced an AH1N2 reassortant virus similar to the A/Lyon/0838/2003 (H1N2) virus. This result provided an experimental evidence of the compatibility between their respective surface H1 and N2 glycoproteins, and suggests that similar events may occur silently amongst human subtypes.

Abstract: The genome of influenza A viruses is comprised of eight negative-sense viral RNAs (vRNAs) that form viral ribonucleoproteins (vRNPs). In order to be infectious, an influenza A viral particle must encapsidate at least one copy of each of the vRNAs. Thus, even though genome segmentation is evolutionary advantageous, it undeniably complicates viral assembly, which is believed to occur through a selective mechanism that still remains to be understood. Using electron tomography 3D-reconstructions, we show that the eight vRNPs of an influenza A Moscow/10/99 (H3N2) virus are interconnected within a star-like structure as they emerge from a unique “transition zone” at the budding tip of the virions. Notably, this “transition zone” is thick enough to accommodate all described packaging signals. We also report that, in vitro, each vRNA segment is involved in a direct contact with at least one other vRNA partner, in a single network of intermolecular interactions. We show that in several cases, the regions involved in vRNA/vRNA interactions overlap with previously identified packaging signals. Our results thus provide support for the involvement of RNA/RNA interactions in the selection and specific packaging of influenza A genomic RNAs, which appear embedded into an organised supramolecular complex likely held together by direct base-pairings between packaging signals. [Copyright &y& Elsevier]