Yu Li, Takeda Kazuyo, Gao Yamei
Division of Viral Products, Office of Vaccines Research and Review, USA.
Microscopy and Imaging Core Facility, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, Maryland, USA.
Virology. 2017 Jun;506:130-140. doi: 10.1016/j.virol.2017.03.016. Epub 2017 Apr 4.
Flavivirus genome encodes seven non-structural proteins (NSPs) and these NSPs are believed to be involved in their genomic RNA replication, of which the mechanism is unclear. We find that West Nile virus (WNV) NSPs were capable of self-assembling membranous vesicles in cells, which are composed of the host endoplasmic reticulum membrane integrated with viral NS1 and NS4A, and possibly NS2A. The vesicles can further organize into replication complex (RC)-associated vesicles which combine both the vesicle and predicted RC components. The authentic RC-associated vesicles were observed in cells transfected with infectious WNV cDNA as well as WNV replicon. Further mutational analysis showed that WNV/DENV heterologous NS polyproteins derived from lethal chimeric recombinants produced abnormal vesicles. Site-directed mutation of either NS2A or NS4A, which resulted in failure of viral RNA replication, caused immature vesicles too. These findings reveal molecular composition and assembly of the virus-specific nanomachine and confirm that these structures are used for the viral RNA replication.
黄病毒基因组编码七种非结构蛋白（NSPs），据信这些NSPs参与其基因组RNA复制，但其机制尚不清楚。我们发现西尼罗河病毒（WNV）的NSPs能够在细胞中自组装膜性囊泡，这些囊泡由整合了病毒NS1和NS4A以及可能还有NS2A的宿主内质网组成。这些囊泡可进一步组织成与复制复合体（RC）相关的囊泡，其结合了囊泡和预测的RC成分。在用感染性WNV cDNA以及WNV复制子转染的细胞中观察到了真正的与RC相关的囊泡。进一步的突变分析表明，源自致死性嵌合重组体的WNV/登革病毒异源NS多蛋白产生异常囊泡。NS2A或NS4A的定点突变导致病毒RNA复制失败，也会产生不成熟的囊泡。这些发现揭示了病毒特异性纳米机器的分子组成和组装，并证实这些结构用于病毒RNA复制。
