Tani Hideki, Iha Koichiro, Shimojima Masayuki, Fukushi Shuetsu, Taniguchi Satoshi, Yoshikawa Tomoki, Kawaoka Yoshihiro, Nakasone Naoe, Ninomiya Haruaki, Saijo Masayuki, Morikawa Shigeru
Special Pathogens Laboratory, Department of Virology I, National Institute of Infectious Diseases, University of Tokyo, Tokyo, Japan.
Special Pathogens Laboratory, Department of Virology I, National Institute of Infectious Diseases, University of Tokyo, Tokyo, Japan Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo, Japan.
J Virol. 2014 Jul;88(13):7317-30. doi: 10.1128/JVI.00512-14. Epub 2014 Apr 16.
Several arenaviruses are known to cause viral hemorrhagic fever (VHF) in sub-Saharan Africa and South America, where VHF is a major public health and medical concern. The biosafety level 4 categorization of these arenaviruses restricts their use and has impeded biological studies, including therapeutic drug and/or vaccine development. Due to difficulties associated with handling live viruses, pseudotype viruses, which transiently bear arenavirus envelope proteins based on vesicular stomatitis virus (VSV) or retrovirus, have been developed as surrogate virus systems. Here, we report the development of a pseudotype VSV bearing each envelope protein of various species of arenaviruses (AREpv), including the newly identified Lujo virus (LUJV) and Chapare virus. Pseudotype arenaviruses generated in 293T cells exhibited high infectivity in various mammalian cell lines. The infections by New World and Old World AREpv were dependent on their receptors (human transferrin receptor 1 [hTfR1] and α-dystroglycan [αDG], respectively). However, infection by pseudotype VSV bearing the LUJV envelope protein (LUJpv) occurred independently of hTfR1 and αDG, indicating that LUJpv utilizes an unidentified receptor. The pH-dependent endocytosis of AREpv was confirmed by the use of lysosomotropic agents. The fusion of cells expressing these envelope proteins, except for those expressing the LUJV envelope protein, was induced by transient treatment at low pH values. LUJpv infectivity was inhibited by U18666A, a cholesterol transport inhibitor. Furthermore, the infectivity of LUJpv was significantly decreased in the Niemann-Pick C1 (NPC1)-deficient cell line, suggesting the necessity for NPC1 activity for efficient LUJpv infection.
LUJV is a newly identified arenavirus associated with a VHF outbreak in southern Africa. Although cell entry for many arenaviruses has been studied, cell entry for LUJV has not been characterized. In this study, we found that LUJpv utilizes neither αDG nor hTfR1 as a receptor and found unique characteristics of LUJV glycoprotein in membrane fusion and cell entry. Proper exclusion of cholesterol or some kinds of lipids may play important roles in LUJpv cell entry.
已知几种沙粒病毒可在撒哈拉以南非洲和南美洲引发病毒性出血热（VHF），在这些地区，VHF是主要的公共卫生和医学问题。这些沙粒病毒的生物安全4级分类限制了它们的使用，并阻碍了生物学研究，包括治疗药物和/或疫苗的开发。由于处理活病毒存在困难，基于水泡性口炎病毒（VSV）或逆转录病毒短暂携带沙粒病毒包膜蛋白的假型病毒已被开发为替代病毒系统。在此，我们报告了一种携带各种沙粒病毒（AREpv）包膜蛋白的假型VSV的开发，包括新发现的卢乔病毒（LUJV）和查帕雷病毒。在293T细胞中产生的假型沙粒病毒在各种哺乳动物细胞系中表现出高感染性。新世界和旧世界AREpv的感染分别依赖于它们的受体（人转铁蛋白受体1 [hTfR1]和α - dystroglycan [αDG]）。然而，携带LUJV包膜蛋白的假型VSV（LUJpv）的感染独立于hTfR1和αDG发生，这表明LUJpv利用一种未鉴定的受体。通过使用溶酶体促渗剂证实了AREpv的pH依赖性内吞作用。除了表达LUJV包膜蛋白的细胞外，表达这些包膜蛋白的细胞在低pH值下短暂处理可诱导融合。LUJpv的感染性受到胆固醇转运抑制剂U18666A的抑制。此外，在尼曼 - 匹克C1（NPC1）缺陷细胞系中，LUJpv的感染性显著降低，这表明NPC1活性对于LUJpv的有效感染是必需的。
LUJV是一种新发现的与南部非洲VHF疫情相关的沙粒病毒。尽管已经对许多沙粒病毒的细胞进入进行了研究，但尚未对LUJV的细胞进入进行表征。在本研究中，我们发现LUJpv既不利用αDG也不利用hTfR1作为受体，并发现了LUJV糖蛋白在膜融合和细胞进入方面的独特特征。适当排除胆固醇或某些种类的脂质可能在LUJpv的细胞进入中起重要作用。
