title: Influenza A viral nucleoprotein interacts with cytoskeleton scaffolding protein α-actinin-4 for viral replication.
abstract: Influenza A virus (IAV), similar to other viruses, exploits the machinery of human host cells for its survival and replication. We identified α-actinin-4, a host cytoskeletal protein, as an interacting partner of IAV nucleoprotein (NP). We confirmed this interaction using co-immunoprecipitation studies, first in a coupled in vitro transcription-translation assay and then in cells either transiently co-expressing the two proteins or infected with whole IAV. Importantly, the NP-actinin-4 interaction was observed in several IAV subtypes, including the 2009 H1N1 pandemic virus. Moreover, immunofluorescence studies revealed that both NP and actinin-4 co-localized largely around the nucleus and also in the cytoplasmic region of virus-infected A549 cells. Silencing of actinin-4 expression resulted in not only a significant decrease in NP, M2 and NS1 viral protein expression, but also a reduction of both NP mRNA and viral RNA levels, as well as viral titers, 24 h post-infection with IAV, suggesting that actinin-4 was critical for viral replication. Furthermore, actinin-4 depletion reduced the amount of NP localized in the nucleus. Treatment of infected cells with wortmannin, a known inhibitor of actinin-4, led to a decrease in NP mRNA levels and also caused the nuclear retention of NP, further strengthening our previous observations. Taken together, the results of the present study indicate that actinin-4, a novel interacting partner of IAV NP, plays a crucial role in viral replication and this interaction may participate in nuclear localization of NP and/or viral ribonucleoproteins.
authors.affiliation: Virology Group, International Centre for Genetic Engineering & Biotechnology, New Delhi, India.
authors.forename: Shipra
authors.initials: S
authors.lastname: Sharma
authors.forename: Adarsh K
authors.initials: AK
authors.lastname: Mayank
authors.forename: Himani
authors.initials: H
authors.lastname: Nailwal
authors.forename: Shashank
authors.initials: S
authors.lastname: Tripathi
authors.forename: Jenish R
authors.initials: JR
authors.lastname: Patel
authors.forename: John B
authors.initials: JB
authors.lastname: Bowzard
authors.forename: Pratibha
authors.initials: P
authors.lastname: Gaur
authors.forename: Ruben O
authors.initials: RO
authors.lastname: Donis
authors.forename: Jacqueline M
authors.initials: JM
authors.lastname: Katz
authors.forename: Nancy J
authors.initials: NJ
authors.lastname: Cox
authors.forename: Renu B
authors.initials: RB
authors.lastname: Lal
authors.forename: Humaira
authors.initials: H
authors.lastname: Farooqi
authors.forename: Suryaprakash
authors.initials: S
authors.lastname: Sambhara
authors.forename: Sunil K
authors.initials: SK
authors.lastname: Lal
mesh_terms: D000185:Actinin / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D054884:Host-Pathogen Interactions; D006801:Humans; D009980:Influenza A virus / Q000502:physiology*; D019590:Nucleocapsid Proteins; D025941:Protein Interaction Mapping; D021381:Protein Transport; D016601:RNA-Binding Proteins / Q000502:physiology*; D015533:Transcriptional Activation; D014758:Viral Core Proteins / Q000502:physiology*; D014779:Virus Replication
chemical_list: C111453:ACTN4 protein, human; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins; D000185:Actinin
keywords: actinin-4; influenza virus; interaction; localization; nucleoprotein