The immunometabolite itaconate drives S. aureus lung infection

项目摘要/摘要 金黄色葡萄球菌引起社区获得性肺炎和医疗保健相关肺炎，导致肺炎发病率下降 对肺功能和死亡风险的增加，特别是对既有肺部疾病的患者。这些 感染很难根除，因为金黄色葡萄球菌可以转变为适应表型，如持久细胞 生物膜，保护细菌免受宿主吞噬细胞和抗菌因子以及抗生素的侵害。 宿主对病原体免疫反应的一个关键组成部分是产生调节性代谢物，如 衣康酸，在免疫细胞中由免疫反应基因1(IRG1)合成。依塔康酸 平衡宿主免疫细胞中的促炎和抗炎信号，并对细菌施加新陈代谢压力。我们 新近发现衣康酸抑制金黄色葡萄球菌的糖酵解和重建葡萄球菌的代谢 以促进生物膜的形成。伊他康特还限制了关键能源产生途径的使用，这表明 它可能促进耐药的宿存细胞的形成。衣康酸在驱动金黄色葡萄球菌肺中的作用 如果不调查感染对宿主对S。 金星。初步数据显示，IRG1在中性粒细胞中高表达，并与 金黄色葡萄球菌肺部感染时促炎细胞因子的产生增加，这不同于抗金黄色葡萄球菌 衣康酸的炎症效应已在其他细胞类型和感染模型中定义。 这项建议旨在进一步研究衣康酸作为宿主-病原体动态的中心介体。 在金黄色葡萄球菌肺部感染期间，这是金黄色葡萄球菌适应肺部的一个潜在驱动因素。目标1将定义 衣康酸通过代谢组学、吞噬作用调节宿主对金黄色葡萄球菌的免疫应答 分析、氧化爆发分析和scRNA测序以1)确定中性粒细胞是否是主要来源 衣康酸在金黄色葡萄球菌感染中的作用，2)确定衣康酸对中性粒细胞效应功能的影响，以及3) 确定衣康酸在中性粒细胞和其他免疫细胞中差异调节的途径 在体内感染期间的种群。目标2将定义衣康酸在推动金黄色葡萄球菌适应 应用细菌qRT-PCR、三磷酸腺苷定量、抗生素耐受性检测和流式细胞术对1) 确定衣康酸是否驱动转录变化，从而促进体内周围细胞的形成，2)建立 在体外，衣康酸暴露会降低能量代谢，增加抗生素耐受性，3) 量化金黄色葡萄球菌在体内对衣康酸的分裂和生长。总之，这些研究将定义 宿主免疫代谢在金黄色葡萄球菌肺部感染致炎和细菌持久性中的作用。

PROJECT SUMMARY/ABSTRACT Staphylococcus aureus causes community-acquired and healthcare-associated pneumonias that drive a decline in lung function and an increased risk of mortality, especially in patients with preexisting lung disease. These infections are difficult to eradicate because S. aureus can transition to adaptive phenotypes like persister cells and biofilm, which protect the bacteria against host phagocytes and antimicrobial factors, as well as antibiotics. A key component of the host immune response to pathogens is the production of regulatory metabolites like itaconate, which is synthesized in immune cells by the enzyme Immune-Responsive Gene 1 (Irg1). Itaconate balances pro- and anti-inflammatory signaling in host immune cells and exerts metabolic stress on bacteria. We recently demonstrated that itaconate inhibits S. aureus glycolysis and restructures staphylococcal metabolism to promote biofilm formation. Itaconate also limited the use of key energy-producing pathways, suggesting that it may promote the formation of antibiotic-tolerant persister cells. The role of itaconate in driving S. aureus lung infections cannot be completely understood without also investigating its impact on the host response to S. aureus. Preliminary data demonstrate that Irg1 is highly expressed in neutrophils and is associated with increased pro-inflammatory cytokine production during S. aureus lung infection, which differs from the anti- inflammatory effects of itaconate that have been defined in other cell types and infection models. This proposal aims to further investigate itaconate as a central mediator of the host-pathogen dynamic during S. aureus lung infection and a potential driver of S. aureus adaptation to the lung. Aim 1 will define the role of itaconate in regulating the host immune response to S. aureus by using metabolomics, phagocytosis assays, oxidative burst assays, and scRNA-sequencing to 1) determine if neutrophils are a major source of itaconate during S. aureus infection, 2) define the impact of itaconate on neutrophil effector function, and 3) identify the pathways that are differentially regulated by itaconate in neutrophils and other immune cell populations during in vivo infection. Aim 2 will define the role of itaconate in driving S. aureus adaptation to the lung by using bacterial qRT-PCR, ATP quantification, antibiotic tolerance assays, and flow cytometry to 1) determine if itaconate drives transcriptional changes that promote persister cell formation in vivo, 2) establish that itaconate exposure drives reduced energy metabolism and increased antibiotic tolerance ex vivo, and 3) quantify S. aureus division and growth in response to itaconate in vivo. Together, these studies will define the role of host immunometabolism in driving inflammation and bacterial persistence during S. aureus lung infection.