N-Linked Glycosylation Plays an Important Role in Budding of Neuraminidase Protein and Virulence of Influenza Viruses
N-Linked Glycosylation Plays an Important Role in Budding of Neuraminidase Protein and Virulence of Influenza Viruses
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N-连接糖基化在神经氨酸酶蛋白的出芽和流感病毒的毒力中发挥重要作用。
DOI：
10.1128/jvi.02042-20
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发表时间：
2021-02-01
影响因子：
5.4
通讯作者：
Liu, Qinfang
中科院分区：
文献类型：
--
作者：
Bao, Danqi;Xue, Ruixue;Liu, Qinfang
NA是高度糖基化的蛋白质。然而，NLG如何影响NA蛋白的功能仍不清楚。神经氨酸酶（NA）在流感病毒的生命周期中具有多种功能，尤其是在病毒复制的后期。血凝素（HA）和NA都是高度糖基化的蛋白质。HA的N-连接糖基化（NLG）已被报道有助于流感病毒的免疫逃逸和毒力。然而，NA的NLG的功能在很大程度上仍不清楚。在本研究中，我们发现NLG对NA的出芽能力至关重要。衣霉素处理或NLG基因敲除显著抑制NA的出芽。进一步的研究表明，NLG基因敲除在体外和体内都能引起病毒的减毒。值得注意的是，219位的NLG在H1N1流感病毒的出芽、复制和毒力中起重要作用。通过对NLG基因敲除的NA过表达细胞的未折叠蛋白反应（unfolded protein response，UPR）的检测，发现NLG基因敲除的NA主要位于内质网（endoplasmic reticulum，ER），UPR标志物BIP和p-eIF 2 α表达上调，而XBP 1表达下调。所有结果表明，NLG敲除NA堆积在ER中并触发UPR，这可能会关闭NA的萌芽过程。总之，该研究揭示了NA的NLG在病毒复制和出芽中的功能。NA是一种高度糖基化的蛋白质。然而，NLG如何影响NA蛋白的功能仍不清楚。在本研究中，我们发现NLG在NA蛋白的出芽和神经氨酸酶活性中起重要作用。NLG的缺失减弱了病毒出芽和复制。特别是219 NLG位点突变在体内外均能显著减弱H1N1流感病毒的复制和毒力，提示NA蛋白的NLG是流感病毒一种新的毒力标志。
NA is a highly glycosylated protein. Nevertheless, how the NLG affects the function of NA protein remains largely unclear. ABSTRACT Neuraminidase (NA) has multiple functions in the life cycle of influenza virus, especially in the late stage of virus replication. Both of hemagglutinin (HA) and NA are highly glycosylated proteins. N-linked glycosylation (NLG) of HA has been reported to contribute to immune escape and virulence of influenza viruses. However, the function of NLG of NA remains largely unclear. In this study, we found that NLG is critical for budding ability of NA. Tunicamycin treatment or NLG knockout significantly inhibited the budding of NA. Further studies showed that the NLG knockout caused attenuation of virus in vitro and in vivo. Notably, the NLG at 219 position plays an important role in the budding, replication, and virulence of H1N1 influenza virus. To explore the underlying mechanism, the unfolded protein response (UPR) was determined in NLG knockout NA overexpressed cells, which showed that the mutant NA was mainly located in the endoplasmic reticulum (ER), the UPR markers BIP and p-eIF2α were upregulated, and XBP1 was downregulated. All the results indicated that NLG knockout NA was stacked in the ER and triggered UPR, which might shut down the budding process of NA. Overall, the study shed light on the function of NLG of NA in virus replication and budding. IMPORTANCE NA is a highly glycosylated protein. Nevertheless, how the NLG affects the function of NA protein remains largely unclear. In this study, we found that NLG plays important roles in budding and Neuraminidase activity of NA protein. Loss of NLG attenuated viral budding and replication. In particular, the 219 NLG site mutation significantly attenuated the replication and virulence of H1N1 influenza virus in vitro and in vivo, which suggested that NLG of NA protein is a novel virulence marker for influenza viruses.