IRGM proteins as regulators of inflammation

摘要 免疫相关GTPase(IRG)M基因亚型变异与预后不良相关 脓毒症患者；然而，潜在的机制尚不清楚。IRGM和它的老鼠同源基因是 对脂多糖和/或干扰素，特别是巨噬细胞，产生高水平的反应。虽然之前的工作 已经表明，IRGM蛋白结合细胞内膜并调节限制 细胞内病原体的存活，它们在炎症中的主要作用-独立于它们在细胞中的作用- 自主宿主防御--大多未被探索。根据大量初步数据的指示，这项提议 将解决新的假设，即IRGM蛋白直接下调炎症的产生 细胞因子对内毒素的反应，并且至少通过两种不同的机制来实现：(1)通过塑造 调节胞外内毒素感受器TLR4激活的代谢途径，以及(2)通过调节 对摄入的内毒素进行细胞内处理，以限制胞内内毒素传感器Casp11(CASP4)的激活。我们会 测试IRGM蛋白抑制小鼠促炎信号的两种非排他性机制 以及确定IRGM蛋白在脓毒症和抗菌作用中的作用 体内的炎症。AIM1将确定IRGM蛋白对线粒体功能、代谢产物的影响 TLR4激活后的水平、葡萄糖代谢和细胞因子的产生。变更的因果关系 新陈代谢和/或线粒体功能和改变的细胞因子表达将通过 药理和遗传干预。AIM2将探索IRGM蛋白在加工和 感受胞质内毒素并确定IRGM缺陷导致内毒素触发的机制(S) Casp11过度激活。Aim3将确定IRGM蛋白功能的变化如何影响TLR4和/或 体内依赖Casp11的脓毒症。根据我们的发现，IRGM不足会导致过度 体内炎性细胞因子的产生，我们将定义IRGM蛋白不足的后果 临床相关脓毒症小鼠模型的发病率和存活率。新生成的条件性Irgm基因敲除 模型和PAN-Irgm基因敲除小鼠将被用来确定Irgm蛋白和 IRGM异构体的相互依赖功能。总体而言，我们的研究不仅将提供一种机械性的 对IRGM蛋白作为内毒素激活的促炎信号通路负调控因子的认识 但也定义了这些调控过程在确定不同脓毒症结局方面的相关性 子类型。

ABSTRACT Hypomorphic variants of the Immunity Related GTPase (IRG) M gene are associated with poor outcomes for sepsis patients; however, the underlying mechanism is not known. IRGM and its mouse orthologues are produced at high levels in response to LPS and/or interferons, particularly in macrophages. Although prior work has shown that IRGM proteins bind intracellular membranes and modulate immune responses that restrict survival of intracellular pathogens, their primary roles in inflammation - independent of their roles in cell- autonomous host defense - are mostly unexplored. As instructed by extensive preliminary data, this proposal will address the novel hypothesis that IRGM proteins directly downmodulate the production of inflammatory cytokines in response to LPS, and that they do so through at least two distinct mechanisms: (1) by shaping metabolic pathways that regulate activation of the extracellular LPS sensor TLR4, and (2) by modulating intracellular processing of ingested LPS to limit activation of the cytosolic LPS sensor Casp11 (CASP4). We will test these two non-exclusive mechanisms by which IRGM proteins dampen pro-inflammatory signaling in mouse and human macrophages in vitro, as well as define the roles of IRGM proteins in sepsis and antimicrobial inflammation in vivo. Aim1 will determine the impact of IRGM proteins on mitochondrial function, metabolite levels, glucose metabolism, and cytokine production in response to TLR4 activation. Causation between altered metabolism and/or mitochondrial function and altered cytokine expression will be established through pharmacological and genetic interventions. Aim2 will explore the role of IRGM proteins in the processing and sensing of cytoplasmic LPS and define the mechanism(s) by which IRGM deficiencies lead to LPS-triggered Casp11 hyperactivation. Aim3 will determine how changes in IRGM protein function impact TLR4- and /or Casp11-dependent sepsis in vivo. Instructed by our finding that IRGM insufficiency leads to excessive inflammatory cytokine production to LPS in vivo, we will define the consequences of IRGM protein insufficiencies on morbidity and survival in clinically relevant sepsis mouse models. Newly generated conditional Irgm knockout models and a pan-Irgm knockout mouse will be used to determine cell-specific roles for Irgm proteins and the interdependent functions of Irgm isoforms. Collectively, our studies will not only provide a mechanistic understanding of IRGM proteins as negative regulators of LPS-activated proinflammatory signaling pathways but also define the relevance of these regulatory processes in determining the outcome of different sepsis subtypes.