G45R mutation in the nonstructural protein 1 of A/Puerto Rico/8/1934 (H1N1) enhances viral replication independent of ds
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RESEARCH
Open Access
G45R mutation in the nonstructural protein 1 of A/Puerto Rico/8/1934 (H1N1) enhances viral replication independent of dsRNAbinding activity and type I interferon biology Challika Kaewborisuth1,2, Mark Zanin2, Hans Häcker3, Richard J. Webby2 and Porntippa Lekcharoensuk1,4,5*
Abstract Background: The nonstructural protein 1 (NS1) of influenza A viruses can act as a viral replication enhancer by antagonizing type I interferon (IFN) induction and response in infected cells. We previously reported that A/Puerto Rico/8/1934 (H1N1) (PR8) containing the NS1 gene derived from A/swine/IA/15/1930 (H1N1) (IA30) replicated more efficiently than the wild type virus. Here, we identified amino acids in NS1 critical for enhancing viral replication. Methods: To identify a key amino acid in NS1 which can increase the virus replication, growth kinetics of PR8 viruses encoding single mutation in NS1 were compared in A549 cells. NS1 mutant functions were studied using dsRNA-protein pull down, RIG-I mediated IFNβ-promoter activity assays and growth curve analysis in murine lung epithelial type I (Let1) cells. Results: The G45R mutation in the NS1 of PR8 (G45R/NS1) virus is critical for the enhanced viral replication in A549 cells. G45R/NS1 slightly decreased NS1 binding to dsRNA but did not interfere with its suppression of RIG-Imediated type I IFN production. Likewise, replication of G45R/NS1 virus was increased in comparison to wild type virus in both wild type and type I interferon receptor null Let1 cells. Conclusions: The non-conserved amino acid, R45, enhances viral replication which is apparently independent of dsRNA binding and suppression of type I IFN, suggesting a non-characterized function of NS1 for the enhanced viral replication. As G45R/NS1 virus induced the type I IFN induction and response in infected A549 cells, it is also interesting to investigate virus virulence for further studies. Keywords: Influenza A virus, NS1, G45R, dsRNA binding, Type I IFN, Virus replication
Background Influenza A virus has evolved a variety of mechanisms to evade host cell defenses to facilitate efficient transmission [1, 2], replication and virulence [3]. Escape from host innate immune responses allows influenza A virus to proficiently replicate in the cells. The most important * Correspondence: [email protected] 1 Interdisciplinary Graduate Program in Genetic Engineering, The Graduate School, Kasetsart University, Bangkok 10900, Thailand 4 Department of Microbiology and Immunology, Faculty of Veterinary Medicine, Kasetsart University, 50th Ngamwongwan Rd., Chatuchak, Bangkok 10900, Thailand Full list of author information is available at the end of the article
innate immune response against influenza A virus is the interferon (IFN) system [4]. There are three types of IFN families including type I (−α, − β, −ω and -ɛ), type II (−γ) and type III (−λs or IL28/29) [5]. Type I IFNs, especially -α/β, are essential for host defense against viral infection. It is induced following the recognition of viral components or RN
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