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Influenza virus NS1 protein inhibits pre-mRNA splicing and blocks mRNA nucleocytoplasmic transport
1Centro Nacional de Biotecnología (CSIC), Universidad Autónoma de Madrid, Spain.
Abstract:
The influenza virus RNA segment 8 encodes two proteins, NS1 and NS2, by differential splicing. The collinear transcript acts as mRNA for NS1 protein, while the spliced mRNA encodes NS2 protein. The splicing of NS1 mRNA was studied in cells transfected with a recombinant plasmid that has the cDNA of RNA segment 8 cloned under the SV40 late promoter and polyadenylation signals. As described for influenza virus-infected cells, NS1 mRNA was poorly spliced to yield NS2 mRNA. However, inactivation of the NS1 gene, but not the NS2 gene, led to a substantial increase in the splicing efficiency, as shown by the relative accumulations of NS1 and NS2 mRNAs. This effect was not specific for NS1 mRNA, since the splicing of the endogenous SV40 early transcript was altered in such a way that t-Ag mRNA was almost eliminated. These changes in the splicing pattern coincided with a strong inhibition of the mRNA nucleocytoplasmic transport. Both NS1 and NS2 mRNAs were retained in the nucleus of cells expressing NS1 protein, but no effect was observed when only NS2 protein was expressed. Furthermore, other mRNAs tested, such as T-Ag mRNA and the non-spliceable nucleoprotein transcript, were also retained in the nucleus upon expression of NS1 protein, suggesting that it induced a generalized block of mRNA export from the nucleus.
Insights
Influenza virus NS1 protein inhibits its own mRNA splicing and blocks the export of various mRNAs from the nucleus. This NS1 protein-induced nuclear retention affects multiple viral and cellular transcripts.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Influenza virus RNA segment 8 encodes NS1 and NS2 proteins via differential splicing.
- NS1 protein is derived from a collinear transcript, while NS2 protein comes from a spliced mRNA.
- Understanding the regulation of influenza mRNA splicing and export is crucial for viral replication.
Purpose of the Study:
- To investigate the role of influenza virus NS1 protein in mRNA splicing efficiency.
- To determine the effect of NS1 protein expression on mRNA nucleocytoplasmic transport.
- To elucidate the mechanism by which NS1 influences viral and cellular mRNA processing.
Main Methods:
- Transfection of cells with a recombinant plasmid containing influenza RNA segment 8 cDNA.
- Analysis of mRNA splicing patterns by assessing relative mRNA accumulations.
- Evaluation of mRNA nucleocytoplasmic transport by examining nuclear retention of transcripts.
Main Results:
- Inactivation of the NS1 gene significantly increased mRNA splicing efficiency.
- Expression of NS1 protein led to retention of both NS1 and NS2 mRNAs in the nucleus.
- NS1 protein expression caused a generalized block of mRNA export, affecting viral (T-Ag) and cellular transcripts.
Conclusions:
- Influenza virus NS1 protein plays a critical role in regulating its own mRNA splicing.
- NS1 protein actively inhibits mRNA splicing and induces a broad block of mRNA export from the nucleus.
- The findings suggest NS1 protein interferes with host cell mRNA processing and transport, potentially aiding viral replication.