The influenza A virus NS1 protein promotes efficient nuclear export of unspliced viral M1 mRNA

Influenza A virus mRNAs are transcribed by the viral RNA-dependent RNA polymerase in the cell nucleus before being exported to the cytoplasm for translation. Segment 7 produces two major transcripts: an unspliced mRNA that encodes the M1 matrix protein and a spliced transcript that encodes the M2 ion channel. Export of both mRNAs is dependent on the cellular NXF1/TAP pathway but it is unclear how they are recruited to the export machinery or how the intron-containing but unspliced M1 mRNA bypasses the normal quality control checkpoints.
Using fluorescent in situ hybridization to monitor segment 7 mRNA localisation, we found that cytoplasmic accumulation of unspliced M1 mRNA was inefficient in the absence of NS1, both in the context of segment 7 RNPs reconstituted by plasmid transfection and in mutant virus34
infected cells. This effect was independent of any major effect on steady-state levels of segment 7 mRNA or splicing, but corresponded to a ~ 5-fold reduction in the accumulation of M1. A similar defect in intronless HA mRNA nuclear export was seen with an NS1 mutant virus.
Efficient export of M1 mRNA required both an intact NS1 RNA-binding domain and effector domain. Furthermore, while wildtype NS1 interacted with cellular NXF1 and also increased the interaction of segment 7 mRNA with NXF1, mutant NS1 polypeptides unable to promote mRNA export did neither. Thus we propose that NS1 facilitates late viral gene expression by acting as an adaptor between viral mRNAs and the cellular nuclear export machinery to promote their nuclear export.