Phosphoproteomics to Characterize Host Response During Influenza A Virus Infection of Human Macrophages
Phosphoproteomics to Characterize Host Response During Influenza A Virus Infection of Human Macrophages
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DOI：
10.1074/mcp.m116.057984
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发表时间：
2016-10-01
影响因子：
7
通讯作者：
Nyman, Tuula A.
中科院分区：
文献类型：
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作者：
Soderholm, Sandra;Kainov, Denis E.;Nyman, Tuula A.
甲型流感病毒引起人类呼吸道感染，并引起每年的季节性爆发，以及更罕见的可怕流行病。甲型流感病毒对针对病毒的抗病毒治疗迅速产生耐药性，这是目前的主要治疗选择。一种有希望的替代方法是靶向被流感病毒利用的宿主细胞因子。为此，我们表征了甲型流感病毒感染的原代人巨噬细胞的磷酸化蛋白质组，以阐明流感感染后激活的细胞内信号传导途径和关键宿主因子。我们确定了1675磷蛋白，4004磷酸肽和4146非冗余磷酸化位点。1113种蛋白质（66%）的磷酸化在感染后受到调节，突出了这种全局磷酸化蛋白质组学分析在原代细胞中的重要性。值得注意的是，285个已确定的磷酸化位点以前没有在公开的磷酸化数据库中描述，尽管许多已发表的大规模磷酸化蛋白质组研究使用人类和小鼠细胞系。磷酸化蛋白质组数据的系统生物信息学分析表明，参与泛素/蛋白酶体途径的蛋白质（如TRIM 22和TRIM 25）和抗病毒反应（如MAVS）在感染的巨噬细胞中发生了变化。已知在小GTP酶、丝裂原活化蛋白激酶和细胞周期蛋白依赖性激酶信号传导中发挥作用的蛋白质也在感染后通过磷酸化调节。特别是，流感感染有一个重大的影响，大量的细胞周期蛋白依赖性激酶底物的磷酸化概况。使用细胞周期蛋白依赖性激酶抑制剂的功能研究表明，细胞周期蛋白依赖性激酶活性是有效的病毒复制和激活宿主抗病毒反应所必需的。此外，我们发现细胞周期蛋白依赖性激酶抑制剂保护IAV感染的小鼠免于死亡。总之，我们提供了第一个全面的磷酸化蛋白质组表征甲型流感病毒感染的原代人巨噬细胞，并提供证据表明，细胞周期蛋白依赖性激酶代表潜在的治疗目标，更有效地治疗流感感染。
Influenza A viruses cause infections in the human respiratory tract and give rise to annual seasonal outbreaks, as well as more rarely dreaded pandemics. Influenza A viruses become quickly resistant to the virus-directed antiviral treatments, which are the current main treatment options. A promising alternative approach is to target host cell factors that are exploited by influenza viruses. To this end, we characterized the phosphoproteome of influenza A virus infected primary human macrophages to elucidate the intracellular signaling pathways and critical host factors activated upon influenza infection. We identified 1675 phosphoproteins, 4004 phosphopeptides and 4146 nonredundant phosphosites. The phosphorylation of 1113 proteins (66%) was regulated upon infection, highlighting the importance of such global phosphoproteomic profiling in primary cells. Notably, 285 of the identified phosphorylation sites have not been previously described in publicly available phosphorylation databases, despite many published large-scale phosphoproteome studies using human and mouse cell lines. Systematic bioinformatics analysis of the phosphoproteome data indicated that the phosphorylation of proteins involved in the ubiquitin/proteasome pathway (such as TRIM22 and TRIM25) and antiviral responses (such as MAVS) changed in infected macrophages. Proteins known to play roles in small GTPase-, mitogen-activated protein kinase-, and cyclin-dependent kinase-signaling were also regulated by phosphorylation upon infection. In particular, the influenza infection had a major influence on the phosphorylation profiles of a large number of cyclin-dependent kinase substrates. Functional studies using cyclin-dependent kinase inhibitors showed that the cyclin-dependent kinase activity is required for efficient viral replication and for activation of the host antiviral responses. In addition, we show that cyclin-dependent kinase inhibitors protect IAV-infected mice from death. In conclusion, we provide the first comprehensive phosphoproteome characterization of influenza A virus infection in primary human macrophages, and provide evidence that cyclin-dependent kinases represent potential therapeutic targets for more effective treatment of influenza infections.