Matriptase Proteolytically Activates Influenza Virus and Promotes Multicycle Replication in the Human Airway Epithelium
Matriptase Proteolytically Activates Influenza Virus and Promotes Multicycle Replication in the Human Airway Epithelium
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DOI：
10.1128/jvi.03005-12
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发表时间：
2013-04-01
影响因子：
5.4
通讯作者：
Richter, Martin V.
中科院分区：
文献类型：
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作者：
Beaulieu, Alexandre;Gravel, Emilie;Richter, Martin V.
流感病毒不编码任何蛋白酶，必须依靠宿主蛋白酶对其表面血凝素蛋白进行蛋白水解激活，以便与受感染的宿主细胞融合。最近在了解流感病毒血凝素活化的人蛋白酶方面取得的进展导致鉴定出II型跨膜丝氨酸蛋白酶TMPRSS2和TMPRSS4以及人气道胰蛋白酶样蛋白酶的成员；然而，没有一种被证明是唯一负责血凝素裂解的酶。在这项研究中，我们鉴定并表征了基质酶是一种流感病毒激活蛋白酶，能够支持人呼吸道上皮细胞中的多周期病毒复制。使用共聚焦显微镜，我们发现基质酶在人上皮细胞的顶端表面和内体内与血凝素共定位，我们发现蛋白酶的可溶性形式能够在很宽的pH范围内特异性地切割H1病毒的血凝素，但不能从H2和H3病毒中切割血凝素。我们发现，人支气管上皮细胞基质酶的小干扰RNA （siRNA）敲低可显著阻断流感病毒在这些细胞中的复制。最后，我们提供了一种选择性的、缓慢的、紧密结合的基质酶抑制剂，可显著减少H1N1流感病毒（包括2009年大流行病毒）的病毒复制（减少1.5 log）。我们的研究为基质酶的作用建立了一个三管齐下的模型：在细胞外空间以其棚状形式激活进入的病毒，当病毒以其膜结合和/或棚状形式在上皮细胞的顶端表面附着或退出时，以及在病毒融合发生的膜结合形式在核内体内激活。
Influenza viruses do not encode any proteases and must rely on host proteases for the proteolytic activation of their surface hemagglutinin proteins in order to fuse with the infected host cells. Recent progress in the understanding of human proteases responsible for influenza virus hemagglutinin activation has led to the identification of members of the type II transmembrane serine proteases TMPRSS2 and TMPRSS4 and human airway trypsin-like protease; however, none has proved to be the sole enzyme responsible for hemagglutinin cleavage. In this study, we identify and characterize matriptase as an influenza virus-activating protease capable of supporting multicycle viral replication in the human respiratory epithelium. Using confocal microscopy, we found matriptase to colocalize with hemagglutinin at the apical surface of human epithelial cells and within endosomes, and we showed that the soluble form of the protease was able to specifically cleave hemagglutinins from H1 virus, but not from H2 and H3 viruses, in a broad pH range. We showed that small interfering RNA (siRNA) knockdown of matriptase in human bronchial epithelial cells significantly blocked influenza virus replication in these cells. Lastly, we provide a selective, slow, tight-binding inhibitor of matriptase that significantly reduces viral replication (by 1.5 log) of H1N1 influenza virus, including the 2009 pandemic virus. Our study establishes a three-pronged model for the action of matriptase: activation of incoming viruses in the extracellular space in its shed form, upon viral attachment or exit in its membrane-bound and/or shed forms at the apical surface of epithelial cells, and within endosomes by its membrane-bound form where viral fusion takes place.