C3-Dependent Intracellular Killing in Innate Immunity and Bacterial Pathogenesis

摘要 细菌感染仍然是全球发病率和死亡率的主要原因，也是一个严重的公共卫生问题。 由于抗生素耐药性的增加和疫苗的有限。许多最严重的细菌感染 起源于粘膜表面，如肠道、呼吸道或皮肤，然后通过 血液流动。引起粘膜和侵袭性疾病的两种主要人类病原体是革兰氏 阴性肠杆菌(如鼠伤寒沙门氏菌，STM)和革兰阳性A组链球菌 (天然气)。补体系统是宿主抵抗血流感染的先天防御的关键 蛋白质及其活化级联反应，特别是C3的调理作用，与其杀菌活性相结合。 吞噬细胞包括巨噬细胞(MΦ)和中性粒细胞。STM和GAS的致病菌株可以颠覆 吞噬酶体能在体内外MΦ细胞内存活，因此自噬系统 成为病原体生存/杀戮的关键战场。这个项目汇集了两个高度 经验丰富且富有成效的内科科学家研究人员，具有互补的专业知识：革兰氏阴性 细菌致病机制、粘膜免疫和Gram-Gram-Gram偶联的小鼠(MPI、M.Raffatellu)模型 积极的细菌致病机制、先天免疫和细菌-吞噬细胞相互作用(MPI，V.Nizet)。同舟共济 我们最近发现了C3的一种新的、必不可少的细胞内功能：将细菌定位于自噬 在MΦ中杀死细菌的系统--这一发现可能会挑战M-Φ细菌无数研究的结论 在没有活性血清的情况下进行相互作用。此外，我们还发现STM丝氨酸蛋白酶 PgtE、GAS丝氨酸蛋白酶SpyCEP和GAS半胱氨酸蛋白酶SpeB允许各自的病原体 灭活C3并在MΦ细胞内复制。我们的中心假设是细胞内C3依赖 自噬是宿主固有防御的关键，而像STM和GAS这样的入侵病原体能够 抵消这一过程大大增加了它们的致病潜力。最近的数据还表明， 微生物群在皮肤和粘膜补体的产生中起着至关重要的作用，这可能代表着另一种 共生微生物影响侵袭性细菌感染风险的关键因素。在这里，我们建议继续 为了研究细胞内C3在感染过程中的作用，了解这一新的原理的分支 先天免疫对宿主-病原体相互作用和两种最重要的人类感染结局的影响 疾病。我们的方法可能揭示新的毒力基因和宿主免疫途径，这些途径将连接 机制，解决长期存在的知识差距，并导致具有广泛相关性的新的调查途径 对细菌的致病作用，包括潜在的新的治疗靶点和线索。

SUMMARY Bacterial infections remain a leading cause of morbidity and mortality worldwide and a critical public health issue due to increasing antibiotic resistance and limited vaccines. Many of the most consequential bacterial infections originate at mucosal surfaces, such as the gut, respiratory tract or skin, then disseminate to other tissues via the bloodstream. Two preeminent human pathogens causing both mucosal and invasive diseases are Gram- negative Salmonella enterica (e.g., serovar Typhimurium, STm) and Gram-positive group A Streptococcus (GAS). Pivotal to innate host defense against bloodstream infection is the function of complement system proteins and their activation cascades, especially opsonization by C3, coupled with the bactericidal activity of phagocytic cells including macrophages (MΦ) and neutrophils. Pathogenic strains of STm and GAS can subvert phagolysosome function to survive intracellularly in MΦ ex vivo and in vivo, whereupon the autophagy system emerges as a critical battleground for pathogen survival/killing. This project brings together two highly experienced and productive physician-scientist investigators with complementary expertise: Gram-negative bacterial pathogenesis, mucosal immunity and gnotobiotic mouse models (MPI, M. Raffatellu) coupled to Gram- positive bacterial pathogenesis, innate immunity, and bacterial-phagocyte interactions (MPI, V. Nizet). Together we have recently discovered a novel, essential intracellular function of C3: targeting of bacteria to the autophagy system for killing in MΦ – a discovery that may challenge conclusions of countless studies of MΦ-bacterial interactions performed in the absence of active serum. Further, we have discovered that the STm serine protease PgtE, the GAS serine protease SpyCEP, and the GAS cysteine protease SpeB allow the respective pathogens to inactivate C3 and to replicate intracellularly in MΦ. Our central hypothesis is that intracellular C3-dependent autophagy is critical to host innate defense, and that the ability of invasive pathogens such as STm and GAS to counteract this process substantially increases their disease-causing potential. Recent data also indicate the microbiome plays an essential role in skin and mucosal complement production, which may represent another crucial factor by which commensal microbes affect invasive bacterial infection risk. Here we propose to continue to investigate the role of intracellular C3 during infection, to understand the ramifications of this new principle of innate immunity on host-pathogen interactions and the outcome of two of the most important human infectious diseases. Our approaches are likely to reveal new virulence genes and host immune pathways that will connect mechanisms, resolve longstanding knowledge gaps, and lead to new avenues of investigation of broad relevance to bacterial pathogenesis including potential novel therapeutic targets and leads.