Membrane permeabilization by small hydrophobic nonstructural proteins of Japanese encephalitis virus
Membrane permeabilization by small hydrophobic nonstructural proteins of Japanese encephalitis virus
复制标题
DOI：
10.1128/jvi.73.8.6257-6264.1999
复制
发表时间：
1999-08-01
影响因子：
5.4
通讯作者：
Lin, YL
中科院分区：
文献类型：
--
作者：
Chang, YS;Liao, CL;Lin, YL
日本脑炎病毒(JEV)是一种由蚊子传播的黄病毒，感染JEV可引起人类急性脑炎，并在各种类型的培养细胞中引起严重的细胞病变。我们观察到乙型脑炎病毒复制使感染的仓鼠肾(BHK-21)细胞对翻译抑制剂潮霉素B或α-香草碱敏感，而模拟感染的细胞对此不敏感。然而，关于乙型脑炎病毒非结构(NS)蛋白是否与病毒诱导的膜通透性改变有关的信息很少。利用可诱导的EscherichiaCALL系统，我们研究了JEV NS1至NS4的哪些部分能够改变膜通透性。我们发现JEV蛋白NS2B-NS3的过表达能使细菌细胞对潮霉素B具有通透性，而NS1的过表达不能起到这一作用。当单独表达时，NS2B单独，而不是NS3，足以改变细菌的膜通透性。同样，NS4A或NS4B的表达也使细菌对潮霉素B的抑制敏感。结果表明，NS2B对细菌生长的抑制作用最强，其次是NS2A和NS4A，而NS1、NS3和NS4B对载体的影响较小。此外，当报告基因荧光素酶或P-半乳糖苷酶共转染时，瞬时表达NS2A、NS2B和NS4B显著降低了BHK-21细胞中的报告基因活性。综上所述，我们的结果表明，在JEV感染后，这四个疏水的小NS蛋白对宿主细胞膜的通透性具有不同的修饰作用，从而在一定程度上参与了病毒诱导的感染细胞的细胞病变效应。
Infection with Japanese encephalitis virus (JEV), a mosquito-borne flavivirus, may cause acute encephalitis in humans and induce severe cytopathic effects in various types of cultured cells. We observed that JEV replication rendered infected baby hamster kidney (BHK-21) cells sensitive to the translational inhibitor hygromycin B or alpha-sarcine, to which mock-infected cells were insensitive. However, little is known about whether any JEV nonstructural (NS) proteins contribute to virus-induced changes in membrane permeability. Using an inducible Escherichia call system, we investigated which parts of JEV NS1 to NS4 are capable of modifying membrane penetrability. We found that overexpression of NS2B-NS3, the JEV protease, permeabilized bacterial cells to hygromycin B whereas NS1 expression failed to do so. When expressed separately, NS2B alone, but not NS3, was sufficient to alter bacterial membrane permeability. Similarly, expression of NS4A or NS4B also rendered bacteria susceptible to hygromycin B inhibition. Examination of the effect of NS1 to NS4 expression on bacterial growth rate showed that NS2B exhibited the greatest inhibitory capability, followed by a modest repression from NS2A and NS4A, whereas NS1, NS3, and NS4B had only trivial influence with respect to the vector control. Furthermore, when cotransfected with a reporter gene luciferase or P-galactosidase, transient expression of NS2A, NS2B, and NS4B markedly reduced the reporter activity in BHK-21 cells. Together, our results suggest that upon JEV infection, these four small hydrophobic NS proteins have various modification effects on host cell membrane permeability, thereby contributing in part to virus-induced cytopathic effects in infected cells.