Generation and characterization of organ-tropism mutants of Japanese encephalitis virus in vivo and in vitro
Generation and characterization of organ-tropism mutants of Japanese encephalitis virus in vivo and in vitro
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DOI：
10.1006/viro.1996.0457
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发表时间：
1996-09-01
期刊：
影响因子：
3.7
通讯作者：
Chin, C
中科院分区：
文献类型：
--
作者：
Chen, LK;Lin, YL;Chin, C
利用伽玛射线照射，从台湾分离株NT109中产生了一对乙型脑炎病毒(JEV)的强毒株(RP-9)和减毒株(RP-2MS)。这两个变异体在斑块形态、病毒吸附和在BHK-21细胞中的生长特性上存在差异：(I)RP-2MS产生的空斑比RP-9小；(Ii)RP-2MS对宿主细胞的吸附效率较低，但产生的病毒滴度(爆发大小)较高；(Iii)RP-2MS病毒粒子主要积累在细胞内，而RP-9则释放到细胞外。此外，在体外结合实验中，Rp-9的包膜(E)蛋白，而不是Rp-2MS的包膜蛋白，特异性地与来自BHK-21细胞的57 kDa的细胞蛋白结合。当注射到小鼠脑内时，RP-2MS的毒性比RP-9小得多，致死量分别为>；10(7)和0.4个空斑形成单位的50%。此外，在腹腔接种时，它们的器官趋向性不同，RP-2MS的主要靶器官是肝脏，而RP-9的主要靶器官是脑。这些结果表明，RP-2MS具有较低的神经毒性和较小的外周神经侵袭性。对病毒结构蛋白的分子分析发现，RP-9和RP-2MS的病毒结构蛋白只有两个不同之处：一个是E蛋白，在RP-9和RP-2MS中分别是Glu-138和Lys-138；另一个是PrM，在RP-9中是Tyr-43，在RP-2MS中是His-43。由于PRM的N端92个氨基酸被切割，在成熟的JEV病毒粒子中不存在，E蛋白第138位的单一氨基酸变化可能是体外结合试验中突变体之间差异的原因，E蛋白的这种突变，或者可能与PrM突变一起，可能是两个突变体在体外和体内观察到的表型差异的部分原因。(C)1996年学术出版社。
Using gamma-ray irradiation, a pair of virulent (RP-9) and attenuated (RP-2ms) variants of Japanese encephalitis virus (JEV) were generated from a Taiwanese isolate, NT109. The two variants differed in plaque morphology, virus adsorption, and growth properties in BHK-21 cells: (i) RP-2ms produced smaller plaques than RP-9; (ii) RP-2ms adsorbed less efficiently to host cells but yielded a higher virus titer (burst size); and (iii) RP-2ms virions were mostly accumulated intracellularly, whereas RP-9 was released extracellularly. In addition, in an in vitro binding assay, the envelope (E) protein of RP-9, but not that of RP-2ms, bound specifically to a cellular protein of 57-kDa derived from BHK-21 cells. When injected into mice intracerebrally, RP-2ms was much less virulent than RP-9, with 50% lethal doses of >10(7) and 0.4 plaque forming units, respectively. Moreover, when inoculated intraperitoneally, their organ tropism differed in that the main target organ for RP-2ms was liver, whereas that for RP-9 was brain. These results suggest that RP-2ms was less neurovirulent and less neuroinvasive from peripheral routes. Molecular analysis of the virus structural proteins detected only two differences between RP-9 and RP-2ms: one in E protein, Glu-138 in RP-9 and Lys-138 in RP-2ms, and the other in prM, Tyr-43 in RP-9 and His-43 in RP-2ms. Since the N-terminal 92 amino acids of prM are cleaved and not present in mature JEV virions, the single-amino-acid change of the E protein at position 138 may account for the difference between the mutants in the in vitro binding assay, Such mutation in E protein, or perhaps in conjunction with the prM mutation, may be responsible, in part, for the phenotypic differences observed in vitro and in vivo between the two mutants. (C) 1996 Academic Press, Inc.