DDX60L Is an Interferon-Stimulated Gene Product Restricting Hepatitis C Virus Replication in Cell Culture
DDX60L Is an Interferon-Stimulated Gene Product Restricting Hepatitis C Virus Replication in Cell Culture
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DOI：
10.1128/jvi.01297-15
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发表时间：
2015-10-01
影响因子：
5.4
通讯作者：
Lohmann, Volker
中科院分区：
文献类型：
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作者：
Gruenvogel, Oliver;Esser-Nobis, Katharina;Lohmann, Volker
所有主要类型的干扰素（IFN）在体外和体内均能有效抑制丙型肝炎病毒（HCV）的复制。值得注意的是，HCV复制对肝癌细胞系Huh 6中的IFN-γ不敏感，尽管有完整的信号传导途径。我们在Huh 6和Huh-7细胞之间进行转录组分析以鉴定IFN-γ应答的效应基因，从而鉴定DExD/H盒解旋酶DEAD盒多肽60样（DDX 60 L）作为HCV复制的限制因子。DDX 60 L及其同源物DEAD盒多肽60（DDX 60）均在病毒感染和IFN处理后在原代人肝细胞中诱导。然而，完全敲低DDX 60 L增加了Huh-7细胞中的HCV复制，并从II型IFN以及I型和III型IFN治疗中挽救了HCV复制，这表明DDX 60 L是针对HCV的先天免疫应答的重要效应蛋白。相反，我们发现DDX 60 L对甲型肝炎病毒复制没有影响。DDX 60 L蛋白仅在强烈的异位过表达时才可检测到，显示出广泛的细胞质分布，但在这些条件下引起细胞病变效应。DDX 60 L敲低并没有改变干扰素刺激基因（ISG）诱导后，干扰素治疗，但抑制HCV复制后异位表达，这表明它是一个直接效应的先天免疫反应。它最有可能抑制病毒RNA复制，因为我们发现DDX 60 L既不影响HCV亚基因组复制子的翻译或稳定性，也不影响感染性病毒的组装。与DDX 60相似，DDX 60 L对RIG-I依赖的先天免疫激活有中度影响，表明在病毒RNA.IMPORTANCEInterferons感应额外的功能诱导过多的干扰素刺激基因（ISGs），这是我们对病毒感染的第一道防线。此外，IFN已被用于抗病毒治疗，特别是针对人类病原体丙型肝炎病毒（HCV）;尽管如此，它们的作用机制尚未得到很好的理解，因为不同的，重叠的拮抗效应ISG组靶向具有不同生物学特性的病毒。我们的工作确定DDX 60 L作为一种新的因子，抑制HCV的复制。DDX 60 L的表达与其同源物DDX 60的表达类似，但我们的数据表明它具有不同的功能，因为我们发现DDX 60在对抗HCV复制方面没有贡献。天然免疫反应的新组分的鉴定有助于全面理解控制抗病毒防御的复杂机制。
All major types of interferon (IFN) efficiently inhibit hepatitis C virus (HCV) replication in vitro and in vivo. Remarkably, HCV replication is not sensitive to IFN-gamma in the hepatoma cell line Huh6, despite an intact signaling pathway. We performed transcriptome analyses between Huh6 and Huh-7 cells to identify effector genes of the IFN-gamma response and thereby identified the DExD/H box helicase DEAD box polypeptide 60-like (DDX60L) as a restriction factor of HCV replication. DDX60L and its homolog DEAD box polypeptide 60 (DDX60) were both induced upon viral infection and IFN treatment in primary human hepatocytes. However, exclusively DDX60L knockdown increased HCV replication in Huh-7 cells and rescued HCV replication from type II IFN as well as type I and III IFN treatment, suggesting that DDX60L is an important effector protein of the innate immune response against HCV. In contrast, we found no impact of DDX60L on replication of hepatitis A virus. DDX60L protein was detectable only upon strong ectopic overexpression, displayed a broad cytoplasmic distribution, but caused cytopathic effects under these conditions. DDX60L knockdown did not alter interferon-stimulated gene (ISG) induction after IFN treatment but inhibited HCV replication upon ectopic expression, suggesting that it is a direct effector of the innate immune response. It most likely inhibits viral RNA replication, since we found neither impact of DDX60L on translation or stability of HCV subgenomic replicons nor additional impact on assembly of infectious virus. Similar to DDX60, DDX60L had a moderate impact on RIG-I dependent activation of innate immunity, suggesting additional functions in the sensing of viral RNA.IMPORTANCEInterferons induce a plethora of interferon-stimulated genes (ISGs), which are our first line of defense against viral infections. In addition, IFNs have been used in antiviral therapy, in particular against the human pathogen hepatitis C virus (HCV); still, their mechanism of action is not well understood, since diverse, overlapping sets of antagonistic effector ISGs target viruses with different biologies. Our work identifies DDX60L as a novel factor that inhibits replication of HCV. DDX60L expression is regulated similarly to that of its homolog DDX60, but our data suggest that it has distinct functions, since we found no contribution of DDX60 in combatting HCV replication. The identification of novel components of the innate immune response contributes to a comprehensive understanding of the complex mechanisms governing antiviral defense.