IRGM proteins as regulators of inflammation

摘要 免疫相关GT3（IRG）M基因的亚型变体与以下疾病的不良结局相关： 脓毒症患者;然而，其潜在机制尚不清楚。IRGM及其小鼠直系同源物是 在LPS和/或干扰素的作用下，特别是在巨噬细胞中产生高水平的干扰素。虽然先前的工作 已经表明IRGM蛋白结合细胞内膜并调节免疫反应， 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-大多是未开发的。根据大量的初步数据，这项建议 将解决新的假设，IRGM蛋白直接下调炎症反应的产生， 细胞因子，并且它们通过至少两种不同的机制来这样做：（1）通过形成 调节细胞外LPS传感器TLR 4的活化的代谢途径，和（2）通过调节 摄取的LPS的细胞内加工以限制胞质LPS传感器Casp 11（CASP 4）的激活。我们将 测试IRGM蛋白抑制小鼠促炎信号传导的这两种非排他性机制 以及IRGM蛋白在脓毒症和抗微生物感染中的作用。 体内炎症。Aim 1将确定IRGM蛋白对线粒体功能、代谢产物和细胞周期的影响。 水平，葡萄糖代谢和细胞因子的产生响应TLR 4激活。变更之间的因果关系 代谢和/或线粒体功能和改变的细胞因子表达将通过以下方式建立： 药理学和遗传学干预。Aim 2将探索IRGM蛋白在加工中的作用， 检测细胞质LPS，并定义IRGM缺陷导致LPS触发的 Casp 11过度激活。Aim 3将确定IRGM蛋白功能的变化如何影响TLR 4-和/或TLR 4-受体。 体内Casp 11依赖性脓毒症。根据我们的发现，IRGM不足导致过度 在体内炎症细胞因子的产生LPS，我们将定义IRGM蛋白不相容性的后果 对临床相关脓毒症小鼠模型的发病率和存活率的影响。新生成的条件性Irgm敲除 模型和泛Irgm敲除小鼠将用于确定Irgm蛋白的细胞特异性作用， Irgm同种型的相互依赖的功能。总的来说，我们的研究不仅提供了一个机械的 了解IRGM蛋白作为LPS激活的促炎信号通路的负调节因子 而且还定义了这些调节过程在确定不同脓毒症结局中的相关性， 亚型

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.