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


CONCLUSIONS
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.
Virology Unit, Department of Infectious Disease Surveillance and Control, National Institute for Health and Welfare (THL), Mannerheimintie 166, FIN-00300 Helsinki, Finland. krister.melen@thl.fi
Krister
K
Melén
Janne
J
Tynell
Riku
R
Fagerlund
Pascal
P
Roussel
Danièle
D
Hernandez-Verdun
Ilkka
I
Julkunen
D002460:Cell Line; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D004847:Epithelial Cells / Q000737:chemistry / Q000821:virology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D054884:Host-Pathogen Interactions; D006801:Humans; D053122:Influenza A Virus, H3N2 Subtype / Q000502:physiology*; D018613:Microscopy, Confocal; D019913:Nuclear Localization Signals; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D021381:Protein Transport; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics; D013194:Staining and Labeling; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D000097796:Nucleolin
D002868:Chromosomal Proteins, Non-Histone; D049452:Green Fluorescent Proteins; D019913:Nuclear Localization Signals; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; D000097796:Nucleolin; C085820:INS1 protein, influenza virus; C047458:fibrillarin