Processing of Capsid Protein by Cathepsin L Plays a Crucial Role in Replication of Japanese Encephalitis Virus in Neural and Macrophage Cells
Processing of Capsid Protein by Cathepsin L Plays a Crucial Role in Replication of Japanese Encephalitis Virus in Neural and Macrophage Cells
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DOI：
10.1128/jvi.00477-07
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发表时间：
2007-06
影响因子：
5.4
通讯作者：
Y. Mori;T. Yamashita;Yoshinori Tanaka;Yoshimi Tsuda;T. Abe;K. Moriishi;Y. Matsuura
中科院分区：
文献类型：
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作者：
Y. Mori;T. Yamashita;Yoshinori Tanaka;Yoshimi Tsuda;T. Abe;K. Moriishi;Y. Matsuura
黄病毒衣壳蛋白不仅是核衣壳的组成部分，而且在病毒复制过程中起重要作用。在本研究中，我们在日本脑炎病毒（JEV）感染的细胞中发现了一种小衣壳蛋白，但在病毒颗粒中没有发现。小衣壳蛋白显示通过用宿主半胱氨酸蛋白酶组织蛋白酶L加工而产生。体外切割试验表明，组织蛋白酶L切割氨基酸残基Lys 18和Arg 19之间的衣壳蛋白，这是非常保守的蚊媒黄病毒。通过在JEV的感染性cDNA克隆的切割位点引入取代，产生了对组织蛋白酶L切割衣壳蛋白具有抗性的突变JEV。突变型JEV在猴（Vero）、蚊子（C6/36）和猪（PK 15）细胞系中表现出与野生型JEV相似的生长动力学，而突变型JEV在小鼠巨噬细胞（RAW264.7）和神经母细胞瘤（N18）细胞中的复制受损。此外，突变型JEV对小鼠的神经毒力和神经侵袭力均低于野生型JEV。这些结果表明，组织蛋白酶L加工JEV衣壳蛋白在JEV在神经细胞和巨噬细胞中的复制中起着至关重要的作用，从而导致JEV感染的发病机制。
ABSTRACT The flavivirus capsid protein not only is a component of nucleocapsids but also plays a role in viral replication. In this study, we found a small capsid protein in cells infected with Japanese encephalitis virus (JEV) but not in the viral particles. The small capsid protein was shown to be generated by processing with host cysteine protease cathepsin L. An in vitro cleavage assay revealed that cathepsin L cleaves the capsid protein between amino acid residues Lys18 and Arg19, which are well conserved among the mosquito-borne flaviviruses. A mutant JEV resistant to the cleavage of the capsid protein by cathepsin L was generated from an infectious cDNA clone of JEV by introducing a substitution in the cleavage site. The mutant JEV exhibited growth kinetics similar to those of the wild-type JEV in monkey (Vero), mosquito (C6/36), and porcine (PK15) cell lines, whereas replication of the mutant JEV in mouse macrophage (RAW264.7) and neuroblastoma (N18) cells was impaired. Furthermore, the neurovirulence and neuroinvasiveness of the mutant JEV to mice were lower than those of the wild-type JEV. These results suggest that the processing of the JEV capsid protein by cathepsin L plays a crucial role in the replication of JEV in neural and macrophage cells, which leads to the pathogenesis of JEV infection.