The role of tuft cells in norovirus pathogenesis

项目摘要 人类诺如病毒是全球急性胃肠炎的主要原因， 年然而，没有抗病毒药物或疫苗。我们使用与之密切相关的小鼠或鼠诺如病毒 (MNV)作为了解人类诺如病毒如何引起感染和疾病的模型系统。更广泛地说，我们 我还使用MNV作为一种工具，揭示病毒如何与宿主免疫相互作用的新的和基本的方面， 系统和其他肠道微生物，包括细菌和蠕虫。 我们最近发现CD300lf是MNV的受体。我们利用这一发现将簇状细胞鉴定为 MNV的靶细胞。簇状细胞是一种罕见的上皮细胞，可以感知并启动免疫反应。 对肠道蠕虫的反应。簇状细胞在调节肠道炎症、创伤 恢复和几种粘膜癌。 在初步数据中，我们开发了新的簇状细胞缺陷小鼠和CD300lf条件性敲除小鼠， 和基因敲入小鼠来研究簇状细胞在急性和慢性诺如病毒感染期间的作用。我们也 开发了一种体外MNV簇细胞培养系统，使我们能够剖析 诺如病毒、簇状细胞和免疫系统。本提案的目的是确定以下方面的作用： 诺如病毒感染中的簇状细胞，以确定簇状细胞感染的免疫学后果，以及 以发现感染的簇细胞如何逃避CD8+ T细胞的杀伤。 在目标1中，我们将确定病毒感染是否损害簇细胞功能。我们将确定塔夫特的作用 急性和持续性MNV感染的细胞。我们将确定MNV如何在慢性感染过程中杀死簇细胞， 感染如何扰乱簇细胞转录组，以及病毒感染簇细胞是否损害簇细胞的能力， 保护宿主免受蠕虫感染。 在目标2中，我们将确定毛簇细胞作为免疫保护小生境的机制 关于MNV MNV特异性CD8+ T细胞在慢性感染期间忽略受感染的簇细胞，但它们仍然具有功能 并且能够离体检测抗原。我们将通过以下方法确定簇状细胞的嗜性如何使其对CD8+ T细胞产生抗性： 结合新的小鼠品系、病毒遗传学、簇细胞和T细胞共培养以及两种簇细胞的过继转移， 和MNV特异性CD8+ T细胞。特别是，我们将确定是否静止的长寿命簇细胞，一种簇细胞亚型， 是一种新型的慢性感染的免疫豁免储存库。 我们期望这项工作将产生一个详细的了解细胞嗜性调节诺如病毒 传播，病毒感染在II型免疫中的作用，以及诺如病毒逃避免疫的机制 CD8+ T细胞。这对我们破译人类诺如病毒机制的长期目标至关重要 发病机制和开发成功的人诺如病毒疫苗。

Project Summary Human norovirus is the leading cause of acute gastroenteritis globally causing up to 200,000 deaths per year. However, there are no antiviral drugs or vaccines. We use the closely related mouse or murine norovirus (MNV) as a model system to understand how human norovirus causes infection and disease. More broadly, we also use MNV as a tool to uncover novel and fundamental aspects of how viruses interact with the host immune system and other intestinal microbes including bacteria and worms. We recently discovered CD300lf as a receptor of MNV. We leveraged this finding to identify tuft cells as a target cell of MNV in the mouse intestines. Tuft cells are rare epithelial cells that sense and initiate an immune response against intestinal worms. Tuft cells are also important in regulating intestinal inflammation, wound recovery, and several mucosal cancers. In preliminary data, we developed novel tuft cell deficient mice and both CD300lf conditional knockout and knock-in mice to study the role of tuft cells during acute and chronic norovirus infection. W We also developed an in vitro MNV tuft cell culture system that will enable us to dissect the complex interactions between norovirus, tuft cells, and the immune system. The objectives of this proposal are to determine the role of tuft cells in norovirus infection, to determine the immunological consequences of tuft cell infection, and to discover how infected tuft cells evade CD8+ T cell killing. In Aim 1, we will determine if viral infection impairs tuft cell function. We will identify the role of tuft cells in acute and persistent MNV infection. We will determine how MNV kills tuft cells during chronic infection, how infection perturbs the tuft cell transcriptome, and whether virus infection of tuft cells impairs the ability of tuft cells to protect the host against worm infection. In Aim 2, we will determine the mechanism by which tuft cells act as an immunoprivileged niche for MNV. MNV-specific CD8+ T cells ignore infected tuft cells during chronic infection yet they remain functional and able to detect antigen ex vivo. We will identify how tuft cell tropism enables resistance to CD8+ T cells by combining novel mouse lines, viral genetics, tuft cell and T cell co-culture, and adoptive transfer of both tuft cell and MNV-specific CD8+ T cells. In particular, we will determine if quiescent long-lived tuft cells, a tuft cell subtype, are a novel immunoprivileged reservoir for chronic infection. We anticipate this work will generate a detailed understanding about the cell tropism regulating norovirus transmission, the role of viral infection in type II immunity, and the mechanism by which norovirus evades immune CD8+ T cells. This is critical to our long-term goal of deciphering mechanisms of human norovirus pathogenesis and developing a successful human norovirus vaccine.