<br>全面的甲型流感病毒结构探测分析可识别参与病毒基因组复制的功能性 RNA 结构,Computational and Structural Biotechnology Journal

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全面的甲型流感病毒结构探测分析可识别参与病毒基因组复制的功能性 RNA 结构
Computational and Structural Biotechnology Journal ( IF 4.4 ) Pub Date : 2023-10-19 , DOI: 10.1016/j.csbj.2023.10.036
Naoki Takizawa ₁ , Risa Karakida Kawaguchi ₂

Affiliation

甲型流感病毒基因组被分割成 8 个病毒 RNA （vRNA）。已知 vRNA 的二级结构参与病毒增殖过程。已经分析了体外、病毒和纤维素中的综合 vRNA 结构。然而，结构图的分辨率可以通过比较分析和统计建模来提高。构建更高分辨率和可靠的 RNA 结构图可以识别病毒粒子中 vRNA 上未表征的功能结构基序。在这里，我们使用硫酸二甲酯（DMS）-seq 和通过引物延伸（SHAPE）-seq 分析的选择性 2'-羟基酰化的组合，通过稳健的统计分析建立了病毒粒子中 vRNA 二级结构的全局图谱。我们的高分辨率分析确定了第 6 段核苷酸位置 39 – 60 的茎环结构，并进一步验证了第 5 段核苷酸位置 87 – 130 的结构，该结构先前预测会在计算机中形成假结结构。值得注意的是，当细胞感染具有破坏结构突变的重组病毒时，病毒基因组的复制和包装急剧减少。我们的结果提供了有关病毒体内甲型流感病毒基因组结构的全面和高分辨率信息，并证明甲型流感病毒基因组上的功能性 RNA 结构基序与病毒基因组的适当复制和包装有关。

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Comprehensive in virio structure probing analysis of the influenza A virus identifies functional RNA structures involved in viral genome replication

The influenza A virus genome is segmented into eight viral RNAs (vRNA). Secondary structures of vRNA are known to be involved in the viral proliferation process. Comprehensive vRNA structures in vitro, in virio, and in cellulo have been analyzed. However, the resolution of the structure map can be improved by comparative analysis and statistical modeling. Construction of a more high-resolution and reliable RNA structure map can identify uncharacterized functional structure motifs on vRNA in virion. Here, we establish the global map of the vRNA secondary structure in virion using the combination of dimethyl sulfate (DMS)-seq and selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE)-seq with a robust statistical analysis. Our high-resolution analysis identified a stem-loop structure at nucleotide positions 39 – 60 of segment 6 and further validated the structure at nucleotide positions 87 – 130 of segment 5 that was previously predicted to form a pseudoknot structure in silico. Notably, when the cells were infected with recombinant viruses which possess the mutations to disrupt the structure, the replication and packaging of the viral genome were drastically decreased. Our results provide comprehensive and high-resolution information on the influenza A virus genome structures in virion and evidence that the functional RNA structure motifs on the influenza A virus genome are associated with appropriate replication and packaging of the viral genome.

更新日期：2023-10-19

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