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Influenza A viral nucleoprotein interacts with cytoskeleton scaffolding protein α‐actinin‐4 for viral replication
Author(s) -
Sharma Shipra,
Mayank Adarsh K.,
Nailwal Himani,
Tripathi Shashank,
Patel Jenish R.,
Bowzard John B.,
Gaur Pratibha,
Donis Ruben O.,
Katz Jacqueline M.,
Cox Nancy J.,
Lal Renu B.,
Farooqi Humaira,
Sambhara Suryaprakash,
Lal Sunil K.
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12828
Subject(s) - nucleoprotein , ribonucleoprotein , viral replication , biology , microbiology and biotechnology , influenza a virus , viral matrix protein , viral protein , rna , immunoprecipitation , virology , virus , cell culture , gene , genetics
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 m RNA 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 m RNA 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. Structured digital abstract • NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( 1 , 2 )• NP and actnin-4 colocalize by fluorescence microscopy ( View interaction )• NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( View interaction )• NP binds to actinin-4 by anti tag coimmunoprecipitation ( 1 , 2 )• NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( View interaction )• NP physically interacts with actinin-4 by anti tag coimmunoprecipitation ( View interaction )• NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( View interaction ) • NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( View interaction ) • NP physically interacts with actinin-4 by two hybrid ( 1 , 2 ) • NP physically interacts with actinin-4 by anti bait coimmunoprecipitation ( View interaction )