Mechanisms of immune evasion by a neuroinvasive fungal pathogen

项目摘要 新生隐球菌是一种神经侵袭性酵母，在免疫功能低下的患者中会导致致命性脑膜炎 每年导致18万人死亡，占全球艾滋病相关死亡人数的15%。隐球菌属 在肺部发起感染，在那里它们必须克服宿主的先天免疫来建立复制的利基 并传播到中枢神经系统。Dectin-1是一种重要的模式识别受体(PRR) 启动对病原体相关分子模式β-葡聚糖的抗真菌防御 在真菌细胞壁中无处不在。虽然Dectin-1对于防御重要病原体是必不可少的，例如 在白念珠菌和烟曲霉菌中，该受体不能诱导C。 新生杆菌感染，尽管隐球菌细胞壁中存在β-葡聚糖。这表明 隐球菌使用尚未发现的机制来逃避Dectin-1的感知。这项提议的目标是 来定义这些机制。为了实现这一点，我将检验新生隐孢子虫逃避的中心假设。 Dectin-1利用毒力因子保护PAMP和/或抑制宿主炎症信号的Dectin-1传感。 具体地说，我的目标是1)识别逃避Dectin-1所必需的真菌基因，2)确定相对的 PAMP的屏蔽和炎症抑制在这一过程中的作用。马德哈尼的前期工作 实验室已经建立了一个经过充分验证的、单基因缺失的&gt；4000新城疫杆菌菌株的文库，我有 开发了一个强大的基于测序的工作流程来量化复杂池中的这些突变体。利用这些优势 工具，我将使用活体小鼠系统地识别逃避Dectin-1所需的真菌毒力因子 肺部感染模型和体外巨噬细胞刺激模型。然后定义装饰的机制-- 1逃避，我将分析Dectin-1逃避突变体在PAMP中的不同表型诊断作用 屏蔽或抑制炎症。这将把新的逃避因素归类为通过一种或两种方式起作用 这些机制，从而揭示了PAMP的屏蔽和炎症抑制的程度 影响Dectin-1逃避。因此，通过确定颠覆关键的抗真菌感知途径的因素和 通过定义这一途径被中和的机制，这项工作解决了一个重要的知识 隐球菌发病机制的差距，将为侵袭性真菌如何克服 与生俱来的防御。

Project Summary Cryptococcus neoformans is a neuroinvasive yeast that causes fatal meningitis in immunocompromised individuals, resulting in >180,000 annual deaths representing 15% of global AIDS-related mortality. Cryptococci initiate infection in the lungs, where they must overcome host innate immunity to establish a replicative niche and disseminate to the central nervous system. Dectin-1 is a critical pattern recognition receptor (PRR) that initiates antifungal defenses in response to β-glucans, a pathogen-associated molecular pattern (PAMP) ubiquitous in fungal cell walls. While Dectin-1 is essential for defense against important pathogens such as Candida albicans and Aspergillus fumigatus, this receptor fails induce protective responses during C. neoformans infection, despite the presence of β-glucans in the cryptococcal cell wall. This suggests that cryptococci employ yet-undiscovered mechanisms to evade Dectin-1 sensing. The objective of this proposal is to define these mechanisms. To accomplish this, I will test the central hypothesis that C. neoformans evades Dectin-1 sensing using virulence factors that shield PAMPs and/or suppress host inflammatory signaling. Specifically, I aim to 1) identify fungal genes necessary for Dectin-1 evasion and 2) determine the relative contributions of PAMP shielding and inflammatory suppression to this process. Preliminary work in the Madhani lab has produced a library of >4000 well-validated, single-gene C. neoformans deletion strains, and I have developed a robust sequencing-based workflow to quantify these mutants in complex pools. Leveraging these tools, I will systematically identify fungal virulence factors required for Dectin-1 evasion using an in vivo murine pulmonary infection model and an in vitro macrophage stimulation model. To then define mechanisms of Dectin- 1 evasion, I will profile Dectin-1 evasion mutants for diverse phenotypes diagnostic of roles in either PAMP shielding or inflammatory suppression. This will classify novel evasion factors as acting through one or both of these mechanisms, thereby revealing the extent to which PAMP shielding and inflammatory suppression influence Dectin-1 evasion. Thus, by identifying the factors that subvert a key antifungal sensing pathway and defining the mechanisms through which this pathway is neutralized, this work addresses an important knowledge gap in cryptococcal pathogenesis and will provide a foundational understanding for how invasive fungi overcome innate defenses.