The membrane on the surface of hepatitis E virus particles is derived from the intracellular membrane and contains trans-Golgi network protein 2
The membrane on the surface of hepatitis E virus particles is derived from the intracellular membrane and contains trans-Golgi network protein 2
复制标题
DOI：
10.1007/s00705-013-1912-3
复制
发表时间：
2014-05-01
影响因子：
2.7
通讯作者：
Okamoto, Hiroaki
中科院分区：
文献类型：
--
作者：
Nagashima, Shigeo;Takahashi, Masaharu;Okamoto, Hiroaki
我们以前的研究表明，戊型肝炎病毒（HEV）需要多泡体（MVB）途径释放病毒颗粒，这表明HEV利用细胞质中的细胞ESCRT机制，而不是在细胞表面，从感染的细胞中释放。在这项研究中，我们产生了鼠单克隆抗体（mAb）对膜相关的HEV颗粒，以检查是否膜是来自细胞内囊泡或细胞表面。在免疫捕获RT-PCR检测中，发现已建立的mAb TA 1708捕获膜相关的HEV颗粒，但不捕获膜解离颗粒或粪便HEV。此外，毛地黄皂苷处理证实了细胞培养物产生的HEV颗粒表面上的膜是脂质膜。双免疫荧光染色显示，mAb TA 1708特异性识别trans-Golgi网络蛋白2（TGOLN 2），一种源自trans-Golgi网络的细胞内抗原。支持这些发现的是，在戊型肝炎病毒感染细胞的裂解液中发现了大量表面具有脂膜和ORF 3蛋白的戊型肝炎病毒颗粒。这些结果表明，HEV在细胞质中形成膜相关颗粒，最有可能是通过出芽进入细胞内囊泡，并且释放的具有脂质膜的HEV颗粒在其表面上保留TGOLN 2的抗原性。
Our previous studies demonstrated that hepatitis E virus (HEV) requires the multivesicular body (MVB) pathway to release virus particles, suggesting that HEV utilizes the cellular ESCRT machinery in the cytoplasm, not at the cell surface, to be released from infected cells. In this study, we generated a murine monoclonal antibody (mAb) against the membrane-associated HEV particles to examine whether the membrane is derived from intracellular vesicles or the cell surface. An established mAb, TA1708, was found to capture the membrane-associated HEV particles, but not the membrane-dissociated particles or fecal HEV, in an immunocapture RT-PCR assay. Furthermore, digitonin treatment confirmed that the membrane on the surface of cell-culture-generated HEV particles was a lipid membrane. Double immunofluorescence staining revealed that mAb TA1708 specifically recognizes trans-Golgi network protein 2 (TGOLN2), an intracellular antigen derived from the trans-Golgi network. Supporting these findings, HEV particles with lipid membranes and ORF3 proteins on their surface were found abundantly in the lysates of HEV-infected cells. These results indicate that HEV forms membrane-associated particles in the cytoplasm, most likely by budding into intracellular vesicles, and that the released HEV particles with a lipid membrane retain the antigenicity of TGOLN2 on their surface.