The small envelope protein of porcine reproductive and respiratory syndrome virus possesses ion channel protein-like properties.
The small envelope protein of porcine reproductive and respiratory syndrome virus possesses ion channel protein-like properties.
复制标题
DOI：
10.1016/j.virol.2006.07.013
复制
发表时间：
2006-11-10
期刊：
影响因子：
3.7
通讯作者：
Yoo D
中科院分区：
文献类型：
--
作者：
Lee C;Yoo D
猪繁殖与呼吸综合征病毒(PRRSV)小囊膜蛋白(E蛋白)是编码于mRNA2双顺反子内部开放阅读框(ORF)的73个氨基酸的疏水性蛋白。作为了解E蛋白在PRRSV复制过程中生物学作用的第一步，E基因在全长感染性克隆中的表达被阻断，方法是将ATG翻译起始突变为GTG，从而使全长突变的基因组克隆无法合成E蛋白。用E基因敲除的基因组克隆对PRRSV敏感细胞进行DNA转染，结果显示病毒不具有感染性。而P129-ΔE转基因细胞则在培养上清液中产生病毒颗粒，这些颗粒含有病毒基因组核酸，表明E蛋白是PRRSV感染所必需的，但对病毒颗粒的组装是不可缺少的。电子显微镜显示，在没有E的情况下组装的P129-ΔE病毒粒子具有与野生型粒子相似的外观。链特异性逆转录聚合酶链式反应表明，E蛋白阴性、非传染性的P129-ΔE病毒颗粒能够进入细胞，但进一步的复制步骤被中断。PRRSV的进入被认为是通过受体介导的内吞作用，溶血型碱性化合物和已知的离子通道阻滞剂在去涂层过程中都有效地抑制了PRRSV的复制。E蛋白在大肠杆菌介导的细胞生长抑制和膜通透性增加中的表达。在感染PRRSV的细胞和转E基因的细胞中进行的交联实验表明，E蛋白能够形成同源寡聚体。综上所述，我们的数据表明PRRSV E蛋白可能是一种嵌入在病毒包膜中的离子通道蛋白，有助于病毒的剥离和基因组在细胞质中的释放。
The small envelope (E) protein of porcine reproductive and respiratory syndrome virus (PRRSV) is a hydrophobic 73 amino acid protein encoded in the internal open reading frame (ORF) of the bicistronic mRNA2. As a first step towards understanding the biological role of E protein during PRRSV replication, E gene expression was blocked in a full-length infectious clone by mutating the ATG translational initiation to GTG, such that the full-length mutant genomic clone was unable to synthesize the E protein. DNA transfection of PRRSV-susceptible cells with the E gene knocked-out genomic clone showed the absence of virus infectivity. P129-ΔE-transfected cells however produced virion particles in the culture supernatant, and these particles contained viral genomic RNA, demonstrating that the E protein is essential for PRRSV infection but dispensable for virion assembly. Electron microscopy suggests that the P129-ΔE virions assembled in the absence of E had a similar appearance to the wild-type particles. Strand-specific RT-PCR demonstrated that the E protein-negative, non-infectious P129-ΔE virus particles were able to enter cells but further steps of replication were interrupted. The entry of PRRSV has been suggested to be via receptor-mediated endocytosis, and lysomotropic basic compounds and known ion-channel blocking agents both inhibited PRRSV replication effectively during the uncoating process. The expression of E protein in Escherichia coli-mediated cell growth arrests and increased the membrane permeability. Cross-linking experiments in cells infected with PRRSV or transfected with E gene showed that the E protein was able to form homo-oligomers. Taken together, our data suggest that the PRRSV E protein is likely an ion-channel protein embedded in the viral envelope and facilitates uncoating of virus and release of the genome in the cytoplasm.
登录
查看更多内容
影响因子：
1.5
作者：
BENFIELD, DA;NELSON, E;CHLADEK, D
通讯作者：
CHLADEK, D
影响因子：
5.4
作者：
ARROYO, J;BOCETA, M;CARRASCO, L
通讯作者：
CARRASCO, L
影响因子：
3.8
作者：
Lee, C;Yoo, D
通讯作者：
Yoo, D
影响因子：
5.4
作者：
Chang, YS;Liao, CL;Lin, YL
通讯作者：
Lin, YL
影响因子：
3.7
作者：
Lee, C;Calvert, JG;Yoo, D
通讯作者：
Yoo, D