Genetic Analysis of Serum-Derived Defective Hepatitis C Virus Genomes Revealed Novel Viral cis Elements for Virus Replication and Assembly.
Genetic Analysis of Serum-Derived Defective Hepatitis C Virus Genomes Revealed Novel Viral cis Elements for Virus Replication and Assembly.
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血清来源的缺陷丙型肝炎病毒基因组的遗传分析揭示了用于病毒复制和组装的新型病毒顺式元件。
DOI：
10.1128/jvi.02182-17
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发表时间：
2018-04-01
影响因子：
5.4
通讯作者：
Zhong, Jin
中科院分区：
文献类型：
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作者：
Li, Qingchao;Tong, Yimin;Zhong, Jin
丙型肝炎病毒（HCV）存在缺陷病毒基因组（DVG），但其生物学意义尚未得到深入研究。在这里，我们分析了在患者血清中循环的HCV DVG，其结构蛋白编码区具有缺失。在41株HCV临床分离株中，约30%的分离株具有来自同一患者全长基因组的DVG。DVG、病毒血症和丙氨酸氨基转移酶（ALT）水平之间没有相关性，DVG的测序分析显示存在缺失热点，上游位点在E1，下游位点在E2和NS 2。有趣的是，NS 2的核心蛋白和C-末端蛋白酶结构域的编码序列在DVG中总是完整的，尽管这两种蛋白质对于HCV基因组复制都是不稳定的。机制研究表明，跨膜段3（TMS 3）的NS 2，位于其蛋白酶结构域的上游，是必需的切割NS 2-NS 3和复制的DVG。此外，我们确定了一个高度保守的二级结构（SL 750）内的核心结构域2编码区，这是至关重要的HCV基因组包装。总之，我们对血清来源的HCV DVG的分析揭示了新的病毒顺式元件，这些元件在病毒复制和组装中起重要作用。此外，尚未在HCV RNA基因组上鉴定出常规的包装信号，并且HCV基因组特异性包裹成感染性颗粒的潜在机制仍有待揭示。在这里，我们确定了新的病毒顺式元件的HCV生命周期的关键，通过确定的遗传约束，定义血清来源的HCV DVG的边界。我们发现，跨膜段3的NS 2，位于其蛋白酶结构域的上游，所需的切割NS 2-NS 3和复制的DVG。我们确定了一个高度保守的二级结构（SL 750）内的核心编码区，这是至关重要的HCV基因组包装。总之，我们对血清来源的HCV DVG的分析揭示了以前意想不到的对HCV复制和形态发生至关重要的新的顺式元件。
Defective viral genomes (DVGs) of hepatitis C virus (HCV) exist, but their biological significances have not been thoroughly investigated. Here, we analyzed HCV DVGs circulating in patient sera that possess deletions in the structural protein-encoding region. About 30% of 41 HCV clinical isolates possess DVGs that originated from the full-length genome in the same patients. No correlation between DVGs, viremia, and alanine aminotransferase (ALT) levels was found. Sequencing analysis of DVGs revealed the existence of deletion hot spots, with upstream sites in E1 and downstream sites in E2 and NS2. Interestingly, the coding sequences for the core protein and the C-terminal protease domain of NS2 were always intact in DVGs despite the fact that both proteins are dispensable for HCV genome replication. Mechanistic studies showed that transmembrane segment 3 (TMS3) of NS2, located immediately upstream of its protease domain, was required for the cleavage of NS2-NS3 and the replication of DVGs. Moreover, we identified a highly conserved secondary structure (SL750) within the core domain 2-coding region that is critical for HCV genome packaging. In summary, our analysis of serum-derived HCV DVGs revealed novel viral cis elements that play important roles in virus replication and assembly.IMPORTANCE HCV DVGs have been identified in vivo and in vitro, but their biogenesis and physiological significances remain elusive. In addition, a conventional packaging signal has not yet been identified on the HCV RNA genome, and mechanisms underlying the specificity in the encapsidation of the HCV genome into infectious particles remain to be uncovered. Here, we identified new viral cis elements critical for the HCV life cycle by determining genetic constraints that define the boundary of serum-derived HCV DVGs. We found that transmembrane segment 3 of NS2, located immediately upstream of its protease domain, was required for the cleavage of NS2-NS3 and the replication of DVGs. We identified a highly conserved secondary structure (SL750) within the core-coding region that is critical for HCV genome packaging. In summary, our analysis of serum-derived HCV DVGs revealed previously unexpected novel cis elements critical for HCV replication and morphogenesis.