Center for the Study of Hepatitis C, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA.
Virology. 2013 Apr 25;439(1):23-33. doi: 10.1016/j.virol.2013.01.019. Epub 2013 Feb 26.
Hepatitis C virus (HCV) contributes substantially to human morbidity and mortality world-wide. The development of HCV genomes expressing heterologous proteins has enhanced the ability to study viral infection, but existing systems have drawbacks. Recombinant viruses often require adaptive mutations to compensate for reduced viral titers, or rely on an artificial genomic organization that uncouples viral protein expression from recombinant gene expression. Here, we sought to exploit the viral polyprotein processing machinery to express heterologous proteins within the context of the HCV polyprotein. We show that HCV genotypes 2a and 1b permit insertion of reporter proteins between NS5A and NS5B with minimal impact on viral fitness. Using this strategy we constructed reporter genomes exhibiting a wide dynamic range, simplifying analysis of HCV infection in primary hepatocytes. Expression of heterologous proteins within the HCV genome offers new opportunities to analyze HCV infection in experimental systems without perturbing functions of individual viral proteins.
丙型肝炎病毒（HCV）在全球范围内导致了大量的人类发病率和死亡率。表达异源蛋白的 HCV 基因组的开发增强了研究病毒感染的能力，但现有的系统存在缺陷。重组病毒通常需要适应性突变来弥补病毒滴度的降低，或者依赖于一种将病毒蛋白表达与重组基因表达解耦的人工基因组组织。在这里，我们试图利用病毒多蛋白加工机制在 HCV 多蛋白的背景下表达异源蛋白。我们表明，HCV 基因型 2a 和 1b 允许在 NS5A 和 NS5B 之间插入报告蛋白，而对病毒适应性的影响最小。利用这种策略，我们构建了报告基因组，表现出很宽的动态范围，简化了对原代肝细胞中 HCV 感染的分析。在 HCV 基因组中表达异源蛋白为在实验系统中分析 HCV 感染提供了新的机会，而不会干扰单个病毒蛋白的功能。
