Dataset Information

Competition between influenza A virus genome segments.

ABSTRACT: Influenza A virus (IAV) contains a segmented negative-strand RNA genome. How IAV balances the replication and transcription of its multiple genome segments is not understood. We developed a dual competition assay based on the co-transfection of firefly or Gaussia luciferase-encoding genome segments together with plasmids encoding IAV polymerase subunits and nucleoprotein. At limiting amounts of polymerase subunits, expression of the firefly luciferase segment was negatively affected by the presence of its Gaussia luciferase counterpart, indicative of competition between reporter genome segments. This competition could be relieved by increasing or decreasing the relative amounts of firefly or Gaussia reporter segment, respectively. The balance between the luciferase expression levels was also affected by the identity of the untranslated regions (UTRs) as well as segment length. In general it appeared that genome segments displaying inherent higher expression levels were more efficient competitors of another segment. When natural genome segments were tested for their ability to suppress reporter gene expression, shorter genome segments generally reduced firefly luciferase expression to a larger extent, with the M and NS segments having the largest effect. The balance between different reporter segments was most dramatically affected by the introduction of UTR panhandle-stabilizing mutations. Furthermore, only reporter genome segments carrying these mutations were able to efficiently compete with the natural genome segments in infected cells. Our data indicate that IAV genome segments compete for available polymerases. Competition is affected by segment length, coding region, and UTRs. This competition is probably most apparent early during infection, when limiting amounts of polymerases are present, and may contribute to the regulation of segment-specific replication and transcription.

SUBMITTER: Widjaja I

PROVIDER: S-EPMC3469491 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Competition between influenza A virus genome segments.

Widjaja Ivy I de Vries Erik E Rottier Peter J M PJ de Haan Cornelis A M CA

PloS one 20121011 10

Influenza A virus (IAV) contains a segmented negative-strand RNA genome. How IAV balances the replication and transcription of its multiple genome segments is not understood. We developed a dual competition assay based on the co-transfection of firefly or Gaussia luciferase-encoding genome segments together with plasmids encoding IAV polymerase subunits and nucleoprotein. At limiting amounts of polymerase subunits, expression of the firefly luciferase segment was negatively affected by the prese ...[more]

PMID: 23071819

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Project description:BACKGROUND:The segmented RNA genome of avian Influenza viruses (AIV) allows genetic reassortment between co-infecting viruses, providing an evolutionary pathway to generate genetic innovation. The genetic diversity (16 haemagglutinin and 9 neuraminidase subtypes) of AIV indicates an extensive reservoir of influenza viruses exists in bird populations, but how frequently subtypes reassort with each other is still unknown. Here we quantify the reassortment patterns among subtypes in the Eurasian avian viral pool by reconstructing the ancestral states of the subtypes as discrete states on time-scaled phylogenies with respect to the internal protein coding segments. We further analyzed how host species, the inferred evolutionary rates and the dN/dS ratio varied among segments and between discrete subtypes, and whether these factors may be associated with inter-subtype reassortment rate. RESULTS:The general patterns of reassortment are similar among five internal segments with the exception of segment 8, encoding the Non-Structural genes, which has a more divergent phylogeny. However, significant variation in rates between subtypes was observed. In particular, hemagglutinin-encoding segments of subtypes H5 to H9 reassort at a lower rate compared to those of H1 to H4, and Neuraminidase-encoding segments of subtypes N1 and N2 reassort less frequently than N3 to N9. Both host species and dN/dS ratio were significantly associated with reassortment rate, while evolutionary rate was not associated. The dN/dS ratio was negatively correlated with reassortment rate, as was the number of negatively selected sites for all segments. CONCLUSIONS:These results indicate that overall selective constraint and host species are both associated with reassortment rate. These results together identify the wild bird population as the major source of new reassortants, rather than domestic poultry. The lower reassortment rates observed for H5N1 and H9N2 may be explained by the large proportion of strains derived from domestic poultry populations. In contrast, the higher rates observed in the H1N1, H3N8 and H4N6 subtypes could be due to their primary origin as infections of wild birds with multiple low pathogenicity strains in the large avian reservoir.

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Competition between influenza A virus genome segments.

Publications

Competition between influenza A virus genome segments.

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