Mechanisms of immune evasion by hepatitis C virus and their role in establishment of viral persistence

据估计，全球有1.7亿人慢性感染丙型肝炎病毒(HCV)，在许多情况下导致严重的肝脏损害。这种高持久率的原因尚不完全清楚，但很明显，先天和获得性免疫反应未能消除病毒具有特别重要的意义。在第一个资助期，tp3分析了丙型肝炎病毒对功能性抗原提呈的干扰以及丙型肝炎病毒逃避干扰素-γ反应，干扰素是介导非细胞T细胞对丙型肝炎病毒反应的中心细胞因子。建立了真实的人类白细胞抗原A2限制性CD8+T细胞反应的免疫学模型，为研究抗原提呈的调控提供了有力的系统。Tp3的数据进一步表明，丙型肝炎病毒在干扰素-γ存在的情况下可以持续存在，并发现了一些新的候选基因，可能参与干扰素-γ对丙型肝炎病毒的应答。在继续这项工作中，我们将分析50个候选基因在细胞群体中差异调控的功能作用，在存在有效的干扰素-γ反应的情况下维持高水平的丙型肝炎病毒复制。经确认的候选人将接受对其行动机制的深入研究。这项工作将揭示干扰素-γ抵抗对丙型肝炎病毒持续存在的贡献，并阐明针对丙型肝炎病毒的干扰素-γ应答和抗病毒效应蛋白的一般调节。第二个项目将解决能够激活先天免疫反应的病毒复制中间体的周转问题。RNA复制产生大量的病毒病原体相关分子模式(PAMP)，如双链RNA，这些模式可被胞质(RIG-I，MDA5)或内体(TLR3)模式识别受体识别。在持续感染期间，病毒PAMP必须经历规则的周转，这特别解释了病毒诱导的膜小泡中包含的双链复制中间产物。然而，病毒PAMP的衰变途径尚未确定。因此，我们的工作旨在揭示病毒复制中间体的处置机制，并阐明其在维持病毒持久性方面的作用。我们首先将重点放在含有病毒RNA的外体的作用上。最近发现，外切体是由含有亚基因组复制子的细胞释放的，并能够激活浆细胞样树突状细胞(PDC)。对病毒诱导的外切体形成的调控将揭示它们在病毒RNA的正常更新和绕过细胞内天然免疫反应激活方面的功能。对引起急性感染的其他嗜肝病毒的外切体分泌和PDC激活的分析将进一步阐明外切体是正链RNA病毒的普遍现象还是与持续性感染有关。

An estimated 170 million people worldwide are chronically infected with hepatitis C virus (HCV), leading in many cases to severe liver damage. The reasons for the high rate of persistence are not fully understood yet, but it is clear that the failure of innate and adaptive immune responses to eliminate the virus is of particular importance. During the first funding period TP3 analyzed interference of HCV with functional antigen presentation and escape of HCV from interferon-γ response, the central cytokine mediating noncytolitic T-cell response to HCV. An immunological model for authentic HLA-A2 restricted CD8+ T-cell response was established, providing a powerful system to study modulation of antigen presentation. The data of TP3 furthermore demonstrated that HCV persistence in presence of IFN-γ can be achieved and identified a number of novel candidate genes potentially involved in IFN-γ response to HCV. In continuation of this work we will analyze the functional role of fifty candidate genes differentially regulated in cell populations maintaining high levels of HCV replication in presence of efficient IFN-γ response. Validated candidates will be subjected to in depth studies on their mechanism of action. This work will reveal the contribution of IFN-γ resistance to HCV persistence and shed light on the general regulation of IFN-γ response and antiviral effector proteins targeting HCV. A second project will address the turnover of viral replication intermediates capable of activating innate immune responses. RNA replication generates huge amounts of viral pathogen associated molecular patterns (PAMPs), like double stranded RNA, which are recognized by cytosolic (RIG-I, MDA5) or endosomal (TLR3) pattern recognition receptors. During persistent infections viral PAMPs must undergo a regular turnover, this accounts particularly for double stranded replication intermediates contained in virus induced membrane vesicles. However, the pathway of decay of viral PAMPs has not been identified yet. Our work therefore aims to uncover the mechanism of disposal of viral replication intermediates and clarify its role in maintaining viral persistence. We will initially focus on the role of exosomes containing viral RNA. Exosomes have recently been found to be released by cells harboring subgenomic replicons and are capable of activating plasmacytoid dendritic cells (pDC). Modulation of virus induced exosome formation will reveal their function regarding regular turnover of viral RNA and circumventing activation of intracellular innate immune responses. Analysis of exosome secretion and pDC activation by other liver-tropic viruses causing acute infections will furthermore elucidate whether exosomes are a general phenomenon of positive strand RNA viruses or linked to persistent infections.