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Influenza A H3N2 subtype virus NS1 protein targets into the nucleus and binds primarily via its C-terminal NLS2/NoLS to nucleolin and fibrillarin

Krister Melén1*, Janne Tynell1, Riku Fagerlund2, Pascal Roussel3, Danièle Hernandez-Verdun4 and Ilkka Julkunen1

Author Affiliations

1 Virology Unit, Department of Infectious Disease Surveillance and Control, National Institute for Health and Welfare (THL), Mannerheimintie 166, FIN-00300, Helsinki, Finland

2 Signaling Systems Laboratory, Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA, 92093, USA

3 Functional Organization of the Nucleolus, RNA Biology-FRE 3402 CNRS, Université Pierre et Marie Curie, 75252, Paris cedex 5, France

4 Nuclei and Cell Cycle, Institut Jacques Monod-UMR 7592 CNRS, Université Paris Diderot, 75205, Paris cedex 13, France

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Virology Journal 2012, 9:167  doi:10.1186/1743-422X-9-167

Published: 21 August 2012

Abstract

Background

Influenza A virus non-structural protein 1 (NS1) is a virulence factor, which is targeted into the cell cytoplasm, nucleus and nucleolus. NS1 is a multi-functional protein that inhibits host cell pre-mRNA processing and counteracts host cell antiviral responses. Previously, we have shown that the NS1 protein of the H3N2 subtype influenza viruses possesses a C-terminal nuclear localization signal (NLS) that also functions as a nucleolar localization signal (NoLS) and targets the protein into the nucleolus.

Results

Here, we show that the NS1 protein of the human H3N2 virus subtype interacts in vitro primarily via its C-terminal NLS2/NoLS and to a minor extent via its N-terminal NLS1 with the nucleolar proteins, nucleolin and fibrillarin. Using chimeric green fluorescence protein (GFP)-NS1 fusion constructs, we show that the nucleolar retention of the NS1 protein is determined by its C-terminal NLS2/NoLS in vivo. Confocal laser microscopy analysis shows that the NS1 protein colocalizes with nucleolin in nucleoplasm and nucleolus and with B23 and fibrillarin in the nucleolus of influenza A/Udorn/72 virus-infected A549 cells. Since some viral proteins contain NoLSs, it is likely that viruses have evolved specific nucleolar functions.

Conclusion

NS1 protein of the human H3N2 virus interacts primarily via the C-terminal NLS2/NoLS and to a minor extent via the N-terminal NLS1 with the main nucleolar proteins, nucleolin, B23 and fibrillarin.

Keywords:
Influenza A virus; NS1 protein; NoLS; Nucleolus; Nucleolin; B23; Fibrillarin