Nucleocapsid protein-dependent assembly of the RNA packaging signal of Middle East respiratory syndrome coronavirus.
Nucleocapsid protein-dependent assembly of the RNA packaging signal of Middle East respiratory syndrome coronavirus.
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DOI：
10.1186/s12929-018-0449-x
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发表时间：
2018-05-24
影响因子：
11
通讯作者：
Chang SC
中科院分区：
文献类型：
--
作者：
Hsin WC;Chang CH;Chang CY;Peng WH;Chien CL;Chang MF;Chang SC
中东呼吸综合征冠状病毒（MERS-CoV）由正义单链RNA基因组和四种结构蛋白组成：刺突蛋白、包膜蛋白、膜蛋白和核壳蛋白。病毒基因组组装成病毒颗粒涉及病毒结构蛋白，并且据信是通过识别病毒基因组的特定序列和RNA结构介导的。通过电镜观察和共表达病毒结构蛋白的检测，确定并建立了一种产生MERS冠状病毒样颗粒（MERS VLP）的培养体系。使用VLP系统，鉴定了跨越MERS-CoV基因组（命名为PS258（19,712 - 19,969）ME）的核苷酸19，712至19，969的258个核苷酸的RNA片段作为包装信号。RNA包装信号组装成MERS VLP依赖于病毒核衣壳蛋白。此外，PS258（19712-19969）ME的45个核苷酸的稳定茎环亚结构与病毒核衣壳蛋白的N-末端结构域和C-末端结构域相互作用。此外，功能性SARS-CoV RNA包装信号未能组装成MERS VLP，这表明RNA基因组的病毒特异性组装。在将携带异源mRNA GFP-PS258（19712-19969）ME的MERS VLP与病毒容许Huh 7细胞孵育后，通过检测GFP表达来鉴定MERS-oV RNA包装信号。MERS VLP系统可以帮助我们了解病毒的感染和形态发生。
Middle East respiratory syndrome coronavirus (MERS-CoV) consists of a positive-sense, single-stranded RNA genome and four structural proteins: the spike, envelope, membrane, and nucleocapsid protein. The assembly of the viral genome into virus particles involves viral structural proteins and is believed to be mediated through recognition of specific sequences and RNA structures of the viral genome. A culture system for the production of MERS coronavirus-like particles (MERS VLPs) was determined and established by electron microscopy and the detection of coexpressed viral structural proteins. Using the VLP system, a 258-nucleotide RNA fragment, which spans nucleotides 19,712 to 19,969 of the MERS-CoV genome (designated PS258(19712–19969)ME), was identified to function as a packaging signal. Assembly of the RNA packaging signal into MERS VLPs is dependent on the viral nucleocapsid protein. In addition, a 45-nucleotide stable stem-loop substructure of the PS258(19712–19969)ME interacted with both the N-terminal domain and the C-terminal domain of the viral nucleocapsid protein. Furthermore, a functional SARS-CoV RNA packaging signal failed to assemble into the MERS VLPs, which indicated virus-specific assembly of the RNA genome. A MERS-oV RNA packaging signal was identified by the detection of GFP expression following an incubation of MERS VLPs carrying the heterologous mRNA GFP-PS258(19712–19969)ME with virus permissive Huh7 cells. The MERS VLP system could help us in understanding virus infection and morphogenesis.
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