Nogales Aitor, Martinez-Sobrido Luis, Topham David J, DeDiego Marta L
Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA.
David H. Smith Center for Vaccine Biology and Immunology, University of Rochester, Rochester, New York, USA
J Virol. 2017 Feb 14;91(5). doi: 10.1128/JVI.01930-16. Print 2017 Mar 1.
Influenza virus NS1 protein is a nonstructural, multifunctional protein that counteracts host innate immune responses, modulating virus pathogenesis. NS1 protein variability in subjects infected with H3N2 influenza A viruses (IAVs) during the 2010/2011 season was analyzed, and amino acid changes in residues 86, 189, and 194 were found. The consequences of these mutations for the NS1-mediated inhibition of IFN responses and the pathogenesis of the virus were evaluated, showing that NS1 mutations D189N and V194I impaired the ability of the NS1 protein to inhibit general gene expression, most probably because these mutations decreased the binding of NS1 to the cleavage and polyadenylation specificity factor 30 (CPSF30). A recombinant A/Puerto Rico/8/34 (PR8) H1N1 virus encoding the H3N2 NS1-D189N protein was slightly attenuated, whereas the virus encoding the H3N2 NS1-V194I protein was further attenuated in mice. The higher attenuation of this virus could not be explained by differences in the ability of the two NS1 proteins to counteract host innate immune responses, indicating that another factor must be responsible. In fact, we showed that the virus encoding the H3N2 NS1-V194I protein demonstrated a temperature-sensitive (ts) phenotype, providing a most likely explanation for the stronger attenuation observed. As far as we know, this is the first description of a mutation in NS1 residue 194 conferring a ts phenotype. These studies are relevant in order to identify new residues important for NS1 functions and in human influenza virus surveillance to assess mutations affecting the pathogenicity of circulating viruses. Influenza viral infections represent a serious public health problem, with influenza virus causing a contagious respiratory disease that is most effectively prevented through vaccination. The multifunctional nonstructural protein 1 (NS1) is the main viral factor counteracting the host antiviral response. Therefore, influenza virus surveillance to identify new mutations in the NS1 protein affecting the pathogenicity of the circulating viruses is highly important. In this work, we evaluated amino acid variability in the NS1 proteins from H3N2 human seasonal viruses and the effect of the mutations on innate immune responses and virus pathogenesis. NS1 mutations D189N and V194I impaired the ability of the NS1 protein to inhibit general gene expression, and recombinant viruses harboring these mutations were attenuated in a mouse model of influenza infection. Interestingly, a virus encoding the H3N2 NS1-V194I protein demonstrated a temperature-sensitive phenotype, further attenuating the virus .
流感病毒NS1蛋白是一种非结构的多功能蛋白，可对抗宿主的先天免疫反应，调节病毒的致病性。分析了2010/2011季节感染H3N2甲型流感病毒（IAV）的受试者中NS1蛋白的变异性，发现第86、189和194位残基存在氨基酸变化。评估了这些突变对NS1介导的干扰素反应抑制和病毒致病性的影响，结果表明NS1突变D189N和V194I损害了NS1蛋白抑制一般基因表达的能力，很可能是因为这些突变降低了NS1与切割及聚腺苷酸化特异性因子30（CPSF30）的结合。编码H3N2 NS1-D189N蛋白的重组A/波多黎各/8/34（PR8）H1N1病毒略有减毒，而编码H3N2 NS1-V194I蛋白的病毒在小鼠中减毒更明显。这种病毒更高的减毒程度无法用两种NS1蛋白对抗宿主先天免疫反应能力的差异来解释，这表明一定有其他因素起作用。事实上，我们发现编码H3N2 NS1-V194I蛋白的病毒表现出温度敏感（ts）表型，这很可能是观察到更强减毒效果的原因。据我们所知，这是首次描述NS1第194位残基的突变赋予ts表型。这些研究对于确定对NS1功能重要的新残基以及在人类流感病毒监测中评估影响流行病毒致病性的突变具有重要意义。流感病毒感染是一个严重的公共卫生问题，流感病毒会引发一种传染性呼吸道疾病，通过接种疫苗可最有效地预防。多功能非结构蛋白1（NS1）是对抗宿主抗病毒反应的主要病毒因子。因此，流感病毒监测以识别NS1蛋白中影响流行病毒致病性的新突变非常重要。在这项工作中，我们评估了H3N2人类季节性病毒NS1蛋白中的氨基酸变异性以及这些突变对先天免疫反应和病毒致病性的影响。NS1突变D189N和V194I损害了NS1蛋白抑制一般基因表达的能力，携带这些突变的重组病毒在流感感染小鼠模型中减毒。有趣的是，编码H3N2 NS1-V194I蛋白的病毒表现出温度敏感表型，进一步使病毒减毒。
