Steacie Institute for Molecular Sciences, National Research Council Canada, Ottawa, Ontario K1A 0R6, Canada.
J Biol Chem. 2010 Aug 13;285(33):25602-12. doi: 10.1074/jbc.M110.135483. Epub 2010 Jun 8.
Hepatitis C virus (HCV) relies on many interactions with host cell proteins for propagation. Successful HCV infection also requires enzymatic activity of host cell enzymes for key post-translational modifications. To identify such enzymes, we have applied activity-based protein profiling to examine the activity of serine hydrolases during HCV replication. Profiling of hydrolases in Huh7 cells replicating HCV identified CES1 (carboxylesterase 1) as a differentially active enzyme. CES1 is an endogenous liver protein involved in processing of triglycerides and cholesterol. We observe that CES1 expression and activity were altered in the presence of HCV. The knockdown of CES1 with siRNA resulted in lower levels of HCV replication, and up-regulation of CES1 was observed to favor HCV propagation, implying an important role for this host cell protein. Experiments in HCV JFH1-infected cells suggest that CES1 facilitates HCV release because less intracellular HCV core protein was observed, whereas HCV titers remained high. CES1 activity was observed to increase the size and density of lipid droplets, which are necessary for the maturation of very low density lipoproteins, one of the likely vehicles for HCV release. In transgenic mice containing human-mouse chimeric livers, HCV infection also correlates with higher levels of endogenous CES1, providing further evidence that CES1 has an important role in HCV propagation.
丙型肝炎病毒（HCV）依赖于许多与宿主细胞蛋白的相互作用来进行复制。HCV 的成功感染还需要宿主细胞酶的酶活性来进行关键的翻译后修饰。为了鉴定这些酶，我们应用基于活性的蛋白质谱分析来研究 HCV 复制过程中丝氨酸水解酶的活性。在复制 HCV 的 Huh7 细胞中进行水解酶谱分析，鉴定出 CES1（羧酸酯酶 1）为差异活性酶。CES1 是一种内源性肝脏蛋白，参与甘油三酯和胆固醇的加工。我们观察到 CES1 的表达和活性在 HCV 存在时发生改变。用 siRNA 敲低 CES1 会导致 HCV 复制水平降低，而 CES1 的上调被观察到有利于 HCV 的增殖，这表明该宿主细胞蛋白具有重要作用。在 HCV JFH1 感染的细胞中的实验表明，CES1 有助于 HCV 的释放，因为观察到细胞内 HCV 核心蛋白减少，而 HCV 滴度仍然很高。CES1 活性被观察到增加了脂质滴的大小和密度，这是极低密度脂蛋白成熟所必需的，极低密度脂蛋白是 HCV 释放的可能载体之一。在含有人-鼠嵌合肝脏的转基因小鼠中，HCV 感染也与内源性 CES1 水平升高相关，进一步证明 CES1 在 HCV 增殖中具有重要作用。
