Interplay of PA-X and NS1 Proteins in Replication and Pathogenesis of a Temperature-Sensitive 2009 Pandemic H1N1 Influenza A Virus

Abstract

Influenza A viruses (IAVs) cause seasonal epidemics and occasional pandemics, representing a serious public health concern. It has been described that one mechanism used by some IAV strains to escape the host innate immune responses and modulate virus pathogenicity involves the ability of the PA-X and NS1 proteins to inhibit the host protein synthesis in infected cells. It was reported that for the 2009 pandemic H1N1 IAV (pH1N1) only the PA-X protein had this inhibiting capability, while the NS1 protein did not. In this work, we have evaluated, for the first time, the combined effect of PA-X- and NS1-mediated inhibition of general gene expression on virus pathogenesis, using a temperature-sensitive, live-attenuated 2009 pandemic H1N1 IAV (pH1N1 LAIV). We found that viruses containing PA-X and NS1 proteins that simultaneously have (PA/NS1) or do not have (PA/NS1) the ability to block host gene expression showed reduced pathogenicity However, a virus where the ability to inhibit host protein expression was switched between PA-X and NS1 (PA/NS1) presented pathogenicity similar to that of a virus containing both wild-type proteins (PA/NS1). Our findings suggest that inhibition of host protein expression is subject to a strict balance, which can determine the successful progression of IAV infection. Importantly, knowledge obtained from our studies could be used for the development of new and more effective vaccine approaches against IAV. Influenza A viruses (IAVs) are one of the most common causes of respiratory infections in humans, resulting in thousands of deaths annually. Furthermore, IAVs can cause unpredictable pandemics of great consequence when viruses not previously circulating in humans are introduced into humans. The defense machinery provided by the host innate immune system limits IAV replication; however, to counteract host antiviral activities, IAVs have developed different inhibition mechanisms, including prevention of host gene expression mediated by the viral PA-X and NS1 proteins. Here, we provide evidence demonstrating that optimal control of host protein synthesis by IAV PA-X and/or NS1 proteins is required for efficient IAV replication in the host. Moreover, we demonstrate the feasibility of genetically controlling the ability of IAV PA-X and NS1 proteins to inhibit host immune responses, providing an approach to develop more effective vaccines to combat disease caused by this important respiratory pathogen.

PA-X: a key regulator of influenza A virus pathogenicity and host immune responses

“…It has been shown that multiple mechanisms are related to host shutoff in IAV-infected cells. However, recent studies have focused on the following three main mechanisms of global inhibition of host protein expression by IAV infection: (1) blockade of cellular mRNA processing and nuclear export by NS1 [16][17][18][19]; (2) degradation of host RNA Pol II by the viral RNA-dependent RNA polymerase RdRp complex (RdRP) [20][21][22]; and (3) wide-spread host mRNA degradation by the PA-X protein [11,[23][24][25][26][27][28][29][30][31]. In this review, we focus mainly on the shutoff activity of PA-X protein.…”

“…When considering the role of NS1 and PA-X in viral pathogenicity and host shutoff, it is very interesting to see the combined effects of them in modulating viral virulence. Therefore, recently, Nogales et al investigated the interplay of PA-X and NS1-mediated host shutoff in viral replication and pathogenesis using a temperature-sensitive 2009 pandemic H1N1 virus [31]. These researchers found that viruses that simultaneously encode PA-X and NS1, or are deficient in both proteins, are highly attenuated in vivo, suggesting that there is a strict balance between NS1 and PA-X proteins during host gene expression regulation, viral replication, and pathogenesis.…”

“…Thus, these studies clearly suggest that the effects of PA-X on viral replication and virulence are strain or host specific. Considering the concerted role for PA-X and NS1 in blockading host gene expression [28,31], we envisage that the precise impact of PA-X on viral growth and -, [11] Increase in mice [11] Increase in mice [11] -, [11] Increase in mice [11] H5N1(avian) Increase in various mammalian and avian cells, and in mice, chickens, ducks [24,35,84] Increase in 293T, CEF, DEF cells [24,35,84] Increase in CEF, DEF, MDCK and A549 cells [24,35,84] Increase in chickens, ducks and mice [24,35,84] Increase in MDCK cells [35,84) and A549 cells (35] Increase in mice, chickens, ducks [24,35,84] H1N1(2009 pdm) Increase in A549 cells and mice [35]; decrease in Calu-3 cells and mice [33]; decrease in MDCK, A549 and mice [34] Increase in 293T cells [35]; decrease in 293T cells [34] Increase in A549 cells [33][34][35] Increase in mice [35] Increase in A549 cells [35] Increase in mice [33,35]; decrease in mice [34] H9N2 (avian) Have no effect in MDCK and A549 cells, decrease in mice [36] -, [36] Decrease in A549 cells…”

Host Single Nucleotide Polymorphisms Modulating Influenza A Virus Disease in Humans

“…NS1 is a multifunctional protein and a key viral factor that counteracts the host antiviral responses. NS1 has been shown to inhibit the production of interferon (IFN), the activity and expression of multiple interferon-induced genes (ISG) and the processing and nuclear transport of host mRNAs causing cellular shut-off [25,26]. Segment 3 of IAV also encodes two proteins, the polymerase component PA and PA-X.…”

“…Synergistically with NS1, PA-X is also able to block the cellular antiviral responses by inhibiting host protein expression. Moreover, the PA-X protein has been shown to modulate host inflammation, immune responses, apoptosis, and virus pathogenesis [25][26][27][28][29][30].…”