Institut de Recherche en Immunologie et en Cancérologie.
J Virol. 2013 Nov;87(21):11704-20. doi: 10.1128/JVI.01474-13. Epub 2013 Aug 28.
Hepatitis C virus (HCV) orchestrates the different stages of its life cycle in time and space through the sequential participation of HCV proteins and cellular machineries; hence, these represent tractable molecular host targets for HCV elimination by combination therapies. We recently identified multifunctional Y-box-binding protein 1 (YB-1 or YBX1) as an interacting partner of NS3/4A protein and HCV genomic RNA that negatively regulates the equilibrium between viral translation/replication and particle production. To identify novel host factors that regulate the production of infectious particles, we elucidated the YB-1 interactome in human hepatoma cells by a quantitative mass spectrometry approach. We identified 71 YB-1-associated proteins that included previously reported HCV regulators DDX3, heterogeneous nuclear RNP A1, and ILF2. Of the potential YB-1 interactors, 26 proteins significantly modulated HCV replication in a gene-silencing screening. Following extensive interaction and functional validation, we identified three YB-1 partners, C1QBP, LARP-1, and IGF2BP2, that redistribute to the surface of core-containing lipid droplets in HCV JFH-1-expressing cells, similarly to YB-1 and DDX6. Importantly, knockdown of these proteins stimulated the release and/or egress of HCV particles without affecting virus assembly, suggesting a functional YB-1 protein complex that negatively regulates virus production. Furthermore, a JFH-1 strain with the NS3 Q221L mutation, which promotes virus production, was less sensitive to this negative regulation, suggesting that this HCV-specific YB-1 protein complex modulates an NS3-dependent step in virus production. Overall, our data support a model in which HCV hijacks host cell machinery containing numerous RNA-binding proteins to control the equilibrium between viral RNA replication and NS3-dependent late steps in particle production.
丙型肝炎病毒（HCV）通过 HCV 蛋白和细胞机制的顺序参与，在时间和空间上协调其生命周期的不同阶段；因此，这些代表了通过联合治疗消除 HCV 的可行的分子宿主靶标。我们最近发现多功能 Y 盒结合蛋白 1（YB-1 或 YBX1）作为 NS3/4A 蛋白和 HCV 基因组 RNA 的相互作用伙伴，负调控病毒翻译/复制和颗粒产生之间的平衡。为了确定调节感染性颗粒产生的新的宿主因子，我们通过定量质谱方法阐明了人肝癌细胞中的 YB-1 相互作用组。我们鉴定了 71 个与 YB-1 相关的蛋白，其中包括先前报道的 HCV 调节剂 DDX3、异质核 RNP A1 和 ILF2。在潜在的 YB-1 相互作用体中，26 种蛋白质在基因沉默筛选中显著调节 HCV 复制。经过广泛的相互作用和功能验证，我们鉴定了三个 YB-1 伴侣，C1QBP、LARP-1 和 IGF2BP2，它们在 HCV JFH-1 表达细胞中重新分布到含有核心的脂滴表面，类似于 YB-1 和 DDX6。重要的是，这些蛋白质的敲低刺激了 HCV 颗粒的释放和/或出芽，而不影响病毒组装，表明存在负调节病毒产生的功能性 YB-1 蛋白复合物。此外，JFH-1 株中 NS3 Q221L 突变促进了病毒产生，对这种负调控的敏感性降低，这表明该 HCV 特异性 YB-1 蛋白复合物调节病毒产生中的 NS3 依赖性步骤。总之，我们的数据支持一种模型，即 HCV 劫持包含许多 RNA 结合蛋白的宿主细胞机制，以控制病毒 RNA 复制和 NS3 依赖性颗粒产生晚期步骤之间的平衡。
