Identification of novel RNA secondary structures within the hepatitis C virus genome reveals a cooperative involvement in genome packaging.
Identification of novel RNA secondary structures within the hepatitis C virus genome reveals a cooperative involvement in genome packaging.
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DOI：
10.1038/srep22952
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发表时间：
2016-03-14
影响因子：
4.6
通讯作者：
Harris M
中科院分区：
文献类型：
--
作者：
Stewart H;Bingham RJ;White SJ;Dykeman EC;Zothner C;Tuplin AK;Stockley PG;Twarock R;Harris M
丙型肝炎病毒（HCV）基因组的特异性包装被假设为由核心-RNA相互作用驱动。为了鉴定参与该过程的病毒基因组区域，我们使用SELEX（通过指数富集的配体系统进化）来鉴定在体外特异性结合核心的RNA适体。这些适体的多个HCV基因组的比较揭示了一个保守的末端环基序内短RNA茎环结构的存在。我们推测，这些基序，以及在HCV基因组中以统计学显著水平存在的子基序与核心蛋白的相互作用可能驱动病毒体组装。我们在HCV感染性分子克隆JFH-1中突变了8个这些预测的基序，从而产生了一系列预测具有改变的RNA二级结构的突变病毒。在只有一个突变结构的病毒中，RNA复制和病毒滴度没有改变。然而，感染性滴度在具有较高数量的突变区域的病毒中降低。因此，这项工作确定了多个新的RNA基序，似乎有助于基因组包装。我们认为，这些结构作为合作包装信号，以驱动特定的RNA在HCV组装。
The specific packaging of the hepatitis C virus (HCV) genome is hypothesised to be driven by Core-RNA interactions. To identify the regions of the viral genome involved in this process, we used SELEX (systematic evolution of ligands by exponential enrichment) to identify RNA aptamers which bind specifically to Core in vitro. Comparison of these aptamers to multiple HCV genomes revealed the presence of a conserved terminal loop motif within short RNA stem-loop structures. We postulated that interactions of these motifs, as well as sub-motifs which were present in HCV genomes at statistically significant levels, with the Core protein may drive virion assembly. We mutated 8 of these predicted motifs within the HCV infectious molecular clone JFH-1, thereby producing a range of mutant viruses predicted to possess altered RNA secondary structures. RNA replication and viral titre were unaltered in viruses possessing only one mutated structure. However, infectivity titres were decreased in viruses possessing a higher number of mutated regions. This work thus identified multiple novel RNA motifs which appear to contribute to genome packaging. We suggest that these structures act as cooperative packaging signals to drive specific RNA encapsidation during HCV assembly.