Genetic Viral and Host Adaptations to Breach Species Barriers of HCV

项目总结 丙型肝炎病毒(丙型肝炎病毒)是一种重要但未被报道的传染病，在71年导致慢性感染 全世界有一百万人。美国疾病控制与预防中心估计，美国每年约有30,000例新的丙型肝炎病例， 每年约有2万人死亡，这使得丙型肝炎病毒的致命性超过了60种其他传染病的总和，包括 爱滋病毒。然而，导致慢性丙型肝炎和终末期肝病的潜在机制 人们对此了解甚少。虽然慢性丙型肝炎现在可以用直接作用的抗病毒药物有效地治疗 (DaaS)，由于治疗成本极高，预防传播的疫苗仍然是高度优先的 (12周课程8万美元)。此外，一旦被感染，个人患肝病的风险仍然很高。 甚至是治疗后。拟议的工作将建立在我们大量研究成果的基础上，以系统地继续进行 分析造成丙型肝炎病毒跨物种传播障碍的机制。 目的1：确定支持丙型肝炎病毒复制的不同物种的已知宿主因子的机制。 丙型肝炎病毒依赖于多种宿主因子在肝细胞中建立复制。我们的初步数据显示 对于所有被测试的类人猿来说，肽基丙酰异构酶A的同源基因，也被称为亲环素A(CypA)，支持 丙型肝炎病毒RNA复制。然而，远亲物种的CypA，如新旧大陆猴子和 老鼠的效率要低得多。我们的目标是机械地定义这个主机和其他主机之间的潜在不兼容性 与丙型肝炎病毒复制机制的病毒编码组件有关的因素。 目的2：描述跨灵长类动物感染丙型肝炎病毒和免疫反应网络的特征 和啮齿动物物种。我们将研究丙型肝炎病毒是否可以在一组新的干细胞来源中感染和复制 来自一系列进化多样性物种的肝细胞样细胞，包括大猩猩，精选的新旧物种 世界上的猴子和啮齿动物。在第二步中，我们将使用高通量单细胞RNA测序来推导出 与丙型肝炎病毒复制相关的物种特异性转录反应网络。 目的3：研究CD302和Cr1L在体内抑制丙型肝炎病毒感染的作用。人类进入因素 天然免疫迟钝的转基因小鼠只支持低水平的病毒复制，这表明是小鼠 限制因素可能会抑制体内的病毒复制。我们的初步数据显示小鼠CD302和 补体成分(3b/4b)受体1样蛋白(Cr1L)在体外限制丙型肝炎病毒的复制。在这里，我们将评估是否有损失- 功能缺失的mCD302和mCr1L在我们的丙型肝炎病毒进入因子敲入小鼠中增加了丙型肝炎病毒的感染。 这项拟议的工作将为了解丙型肝炎病毒的物种取向提供新的见解。普洛斯实验室已经制造了许多 对该领域的开创性贡献，并将在这项重要的工作中得到我们的长期合作伙伴博士的帮助。 Schwartz(Weill Cornell)、Shalek(麻省理工学院)和Pietschmann(德国TWINCORE)。我们的工作将推动这一领域的发展 在发展适合研究丙型肝炎病毒的小动物模型方面取得进展，从而促进丙型肝炎病毒的研究 感染和免疫反应，这是改进治疗和开发疫苗的必要先导。

Project summary Hepatitis C virus (HCV) is an important and underreported infectious disease, causing chronic infection in ~71 million people worldwide. The CDC estimates there are ~30,000 new cases of HCV every year in the US and about 20,000 deaths annually, making HCV more deadly than 60 other infectious diseases combined, including HIV. However, the underlying mechanisms that lead to chronic HCV infection followed by end-stage liver disease are poorly understood. Although chronic hepatitis C can now be effectively treated with direct-acting antivirals (DAAs), a vaccine to prevent transmission remains a high priority due to extremely high treatment costs ($80,000+ for a 12-week course). Furthermore, once infected, individuals remain at high risk for liver disease even post-treatment. The proposed work will build on our substantial research findings to continue systematically analyzing the mechanisms that create barriers to interspecies HCV transmission. Aim 1: Define mechanisms of known host factors from diverse species that support HCV replication. HCV relies on a variety of host factors to establish replication in hepatocytes. Our preliminary data demonstrate that for all great apes tested, orthologs of peptidylprolyl isomerase A, also known as cyclophilin A (CypA), support HCV RNA replication. However, CypA of distantly related species, such as New and Old World monkeys and mice, is far less efficient. We aim to define mechanistically the underlying incompatibility of this and other host factors with the virally encoded components of the HCV replication machinery. Aim 2: Characterize HCV infection and immune response networks following infection across primate and rodent species. We will study whether HCV can infect and replicate in a novel set of stem cell-derived hepatocyte-like cells from a range of evolutionarily diverse species, including great apes, selected New and Old World monkeys and rodents. In a second step, we will use high-throughput single-cell RNA sequencing to derive species-specific transcriptomic response networks associated with HCV replication. Aim 3: Characterize the impact of CD302 and Cr1L on restricting HCV infection in vivo. Human entry factor transgenic mice with blunted innate immunity only support low-level viral replication, suggesting murine restriction factors may suppress viral replication in vivo. Our preliminary data indicate mouse CD302 and complement component (3b/4b) receptor 1-like (Cr1L) limit HCV replication in vitro. Here, we will assess if loss- of-function of mCD302 and mCr1L augments HCV infection in our HCV entry factor knock-in mice. The proposed work will provide new insights into the species tropism of HCV. The Ploss lab has made many seminal contributions to the field and will be aided in this important work by our long-standing collaborators Drs. Schwartz (Weill Cornell), Shalek (MIT) and Pietschmann (Twincore, Germany). Our work will advance the field of HCV research by making progress in the development of small animal models suitable for studying HCV infection and immune responses, a necessary precursor to improving treatment and developing vaccines.