Huang Xingchuan, Dong Wenjuan, Milewska Aleksandra, Golda Anna, Qi Yonghe, Zhu Quan K, Marasco Wayne A, Baric Ralph S, Sims Amy C, Pyrc Krzysztof, Li Wenhui, Sui Jianhua
National Institute of Biological Sciences, Changping, Beijing, China.
National Institute of Biological Sciences, Changping, Beijing, China China Agricultural University Graduate Program, National Institute of Biological Sciences, Beijing, China.
J Virol. 2015 Jul;89(14):7202-13. doi: 10.1128/JVI.00854-15. Epub 2015 Apr 29.
Human coronavirus (hCoV) HKU1 is one of six hCoVs identified to date and the only one with an unidentified cellular receptor. hCoV-HKU1 encodes a hemagglutinin-esterase (HE) protein that is unique to the group a betacoronaviruses (group 2a). The function of HKU1-HE remains largely undetermined. In this study, we examined binding of the S1 domain of hCoV-HKU1 spike to a panel of cells and found that the S1 could specifically bind on the cell surface of a human rhabdomyosarcoma cell line, RD. Pretreatment of RD cells with neuraminidase (NA) and trypsin greatly reduced the binding, suggesting that the binding was mediated by sialic acids on glycoproteins. However, unlike other group 2a CoVs, e.g., hCoV-OC43, for which 9-O-acetylated sialic acid (9-O-Ac-Sia) serves as a receptor determinant, HKU1-S1 bound with neither 9-O-Ac-Sia-containing glycoprotein(s) nor rat and mouse erythrocytes. Nonetheless, the HKU1-HE was similar to OC43-HE, also possessed sialate-O-acetylesterase activity, and acted as a receptor-destroying enzyme (RDE) capable of eliminating the binding of HKU1-S1 to RD cells, whereas the O-acetylesterase-inactive HKU1-HE mutant lost this capacity. Using primary human ciliated airway epithelial (HAE) cell cultures, the only in vitro replication model for hCoV-HKU1 infection, we confirmed that pretreatment of HAE cells with HE but not the enzymatically inactive mutant blocked hCoV-HKU1 infection. These results demonstrate that hCoV-HKU1 exploits O-Ac-Sia as a cellular attachment receptor determinant to initiate the infection of host cells and that its HE protein possesses the corresponding sialate-O-acetylesterase RDE activity.
Human coronaviruses (hCoV) are important human respiratory pathogens. Among the six hCoVs identified to date, only hCoV-HKU1 has no defined cellular receptor. It is also unclear whether hemagglutinin-esterase (HE) protein plays a role in viral entry. In this study, we found that, similarly to other members of the group 2a CoVs, sialic acid moieties on glycoproteins are critical receptor determinants for the hCoV-HKU1 infection. Interestingly, the virus seems to employ a type of sialic acid different from those employed by other group 2a CoVs. In addition, we determined that the HKU1-HE protein is an O-acetylesterase and acts as a receptor-destroying enzyme (RDE) for hCoV-HKU1. This is the first study to demonstrate that hCoV-HKU1 uses certain types of O-acetylated sialic acid residues on glycoproteins to initiate the infection of host cells and that the HKU1-HE protein possesses sialate-O-acetylesterase RDE activity.
人冠状病毒（hCoV）HKU1是迄今鉴定出的六种hCoV之一，也是唯一一种细胞受体尚未明确的病毒。hCoV-HKU1编码一种血凝素酯酶（HE）蛋白，这是β冠状病毒2a组（2a组）所特有的。HKU1-HE的功能在很大程度上仍未确定。在本研究中，我们检测了hCoV-HKU1刺突蛋白S1结构域与一组细胞的结合情况，发现S1可以特异性结合人横纹肌肉瘤细胞系RD的细胞表面。用神经氨酸酶（NA）和胰蛋白酶预处理RD细胞可大大降低这种结合，这表明这种结合是由糖蛋白上的唾液酸介导的。然而，与其他2a组冠状病毒不同，例如hCoV-OC43，其9-O-乙酰化唾液酸（9-O-Ac-Sia）作为受体决定簇，HKU1-S1既不与含9-O-Ac-Sia的糖蛋白结合，也不与大鼠和小鼠红细胞结合。尽管如此，HKU1-HE与OC43-HE相似，也具有唾液酸-O-乙酰酯酶活性，并作为一种受体破坏酶（RDE），能够消除HKU1-S1与RD细胞的结合，而O-乙酰酯酶无活性的HKU1-HE突变体则失去了这种能力。使用人原发性纤毛气道上皮（HAE）细胞培养物，这是hCoV-HKU1感染的唯一体外复制模型，我们证实用HE而不是酶无活性的突变体预处理HAE细胞可阻断hCoV-HKU1感染。这些结果表明，hCoV-HKU1利用O-Ac-Sia作为细胞附着受体决定簇来启动宿主细胞的感染，并且其HE蛋白具有相应的唾液酸-O-乙酰酯酶RDE活性。
人冠状病毒（hCoV）是重要的人类呼吸道病原体。在迄今鉴定出的六种hCoV中，只有hCoV-HKU1没有明确的细胞受体。血凝素酯酶（HE）蛋白在病毒进入过程中是否起作用也不清楚。在本研究中，我们发现，与2a组冠状病毒的其他成员类似，糖蛋白上的唾液酸部分是hCoV-HKU1感染的关键受体决定簇。有趣的是，该病毒似乎使用了一种与其他2a组冠状病毒不同的唾液酸类型。此外，我们确定HKU1-HE蛋白是一种O-乙酰酯酶，并作为hCoV-HKU1的受体破坏酶（RDE）。这是第一项证明hCoV-HKU1利用糖蛋白上某些类型的O-乙酰化唾液酸残基来启动宿主细胞感染并且HKU1-HE蛋白具有唾液酸-O-乙酰酯酶RDE活性的研究。
