The live attenuated Japanese encephalitis virus vaccine SA14-14-2 demonstrated ≥ 95% efficacy and is today the vaccine of choice against JEV globally. Relative to its parent strain SA14, SA14-14-2 carries 46 nucleotide and 24 amino acid alterations, with 8 of the latter located within the envelope glycoprotein. The vaccine strain also fails to synthesize the nonstructural protein NS1’ owing to a silent mutation that abrogates a -1-frameshifting event close to the 5’ end of the NS2A coding sequence. Previous studies employing reverse genetics and mouse models implicated both absence of NS1’ and mutated E, in attenuation of SA14-14-2. We demonstrate progressive reduction in ER stress sensor PERK levels and increased expression of CEBP-homologous protein (CHOP), accompanied by dephosphorylation of eIF2α, inhibition of autophagy maturation and necroptosis following infection of cultured cells with wild-type JEV strain P20778. Autonomous expression of NS1’ caused constitutive up-regulation of CHOP and loss of PERK. Conversely, infection with SA14-14-2 led to significantly increased IRE-1α activation, ER chaperone levels and autophagy. We report labile conformational epitopes accompanied by drastically reduced folding kinetics of intracellular SA14-14-2 envelope protein engendered by sluggish oxidation of cysteine sulfhydryl groups to form disulfide bonds within the endoplasmic reticulum along with altered envelope epitopes in extracellular SA14-14-2 viral particles. We also demonstrate near total conversion of prM to pr and M in SA14-14-2 virus particles. These alterations were accompanied by enhanced activation of mouse and human antigen presenting cells by SA14-14-2 along with superior CD8 + recall T cell responses to viral structural proteins in volunteers vaccinated with SA14-14-2.

中文翻译：

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减毒的日本脑炎病毒减毒活疫苗SA14-14-2减毒的分子机制

减毒的日本脑炎病毒减毒活疫苗SA14-14-2表现出≥95％的功效，如今已成为全球范围内针对JEV的首选疫苗。相对于其亲本菌株SA14，SA14-14-2携带46个核苷酸和24个氨基酸的改变，其中8个位于包膜糖蛋白内。由于沉默突变消除了NS2A编码序列5'末端附近的-1移码事件，该疫苗菌株也不能合成非结构蛋白NS1'。先前采用反向遗传学和小鼠模型的研究表明，SA14-14-2的减毒既不存在NS1'，也不存在突变的E。我们证明了ER应激传感器PERK水平的逐渐降低和CEBP同源蛋白（CHOP）的表达增加，并伴有eIF2α的去磷酸化，抑制野生型JEV株P20778感染培养细胞后自噬成熟和坏死性坏死。NS1'的自主表达引起CHOP的组成型上调和PERK的丢失。相反，感染SA14-14-2导致IRE-1α活化，ER伴侣水平和自噬显着增加。我们报告不稳定的构象表位，伴随着胞内SA14-14-2包膜蛋白的折叠动力学急剧降低，半胱氨酸巯基的缓慢氧化导致内质网内形成二硫键以及胞外SA14-14-2病毒颗粒的包膜表位改变。我们还证明了SA14-14-2病毒颗粒中prM几乎全部转化为pr和M。