The role of Listeria cyclic-di-AMP during infection and immunity

李斯特菌环二腺苷在感染和免疫过程中的作用

• 批准号: 8378115
• 负责人: DANIEL A PORTNOY
• 金额: $ 44.72万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8378115
• 关键词: Address; Adenylate Cyclase; Adjuvant; Bacteria; Biochemistry; Cellular Immunity; Collaborations; Communicable Diseases; Data; Dendritic Cells; Dendritic cell activation; Ethylnitrosourea; Genes; Genetic Screening; Goals; Growth; Immune response; Immunity; In Vitro; Infection; Integration Host Factors; Interferons; Lead; Life; Ligands; Listeria; Listeria monocytogenes; Modeling; Molecular; Mutagenesis; Mycobacterium tuberculosis; Natural Immunity; Pathogenesis; Pathway interactions; Physiological; Production; Role; Signaling Molecule; Testing; efflux pump; macrophage; novel; pathogen; phosphoric diester hydrolase; response; therapeutic vaccine; vaccine development

This proposal is Project 1 within a P01 renewal, entitled "Intracellular pathogens and innate immunity." Listeria monocytogenes is a highly amenable model intracellular pathogen that induces robust and long-lived cell mediated immunity. The goal of the previous proposal was to identify L. monocytogenes determinants that activate a host cytosolic surveillance pathway (CSP) of innate immunity that leads to the expression of IFNB and co-regulated genes. We hypothesize that this pathway contributes significantly to the potent and long-lived cell-mediated immunity induced by L. monocytogenes. Using a genetic screen and subsequent biochemistry, we identified c-di-AMP as the listerial ligand that is secreted through a multidrug efflux pump and activates the CSP. Thus, c-di-AMP is a newly discovered PAMP. C-di-AMP is also a novel and conserved small bacterial signaling molecule that appears essential for bacterial growth. We have identified the di-adenylate cyclase (DacA) and c-di-AMP phosphodiesterase (Pde). In Aim 1, we propose to further characterize the molecular determinants that lead to expression of c-di-AMP and generate a panel of well-characterized strains that express either enhanced or diminished levels of c-di-AMP. We will also address the following questions: What are the bacterial components that regulate c-di-AMP production? Does c-di- AMP act from the inside and/or the outside of bacteria? What is the physiologic role of c-di-AMP? In Aim 2, we will use the strains constructed above to determine the role of c-di-AMP during infection of macrophages and dendritic cells in vitro. We will also evaluate the effects of purified c-di-AMP on dendritic cell activation. In collaboration with the Vance Lab (Project 3), we will use ENU mutagenesis to identify novel host factors that control the CSP. In preliminary data, we identified Sting (Stimulator of interferon genes) as a critical host component necessary for response to L. monocytogenes, M. tuberculosis, and c-di-AMP. In the final Aim, we will test the hypothesis that c-di-AMP production is essential for the full expression of L. monocytogenes pathogenesis, and importantly, contributes to the induction of cell-mediated immunity. We will examine the role of c-di-AMP as an adjuvant.

该提案是P01更新中的项目1，标题为“细胞内病原体和先天免疫”。" 单核细胞增多性李斯特菌是一种高度顺从的细胞内病原体模型，其诱导稳定且长寿命的 细胞免疫先前提案的目标是确定L。单核细胞增多决定簇 其激活先天免疫的宿主胞质监视途径（CSP）， IFNB和共调节基因。我们假设这一途径对有效的和 L.单核细胞增多症使用基因筛选和随后的 在生物化学中，我们鉴定了c-di-AMP作为通过多药外排泵分泌的配体 激活CSP因此，c-di-AMP是一种新发现的PAMP。C-di-AMP也是一种新颖的， 保守的小细菌信号分子，似乎是细菌生长所必需的。我们已经确定 二腺苷酸环化酶（DacA）和c-di-AMP磷酸二酯酶（Pde）。在目标1中，我们建议进一步 表征导致c-di-AMP表达的分子决定因素，并生成一组充分表征的 表达增强或降低水平的c-di-AMP的菌株。我们也要谈谈 以下问题：调节c-di-AMP生产的细菌成分是什么？你... AMP是从细菌的内部和/或外部起作用的？c-di-AMP的生理作用是什么？在目标2中， 我们将使用上述构建的菌株来确定c-di-AMP在巨噬细胞感染过程中的作用 和树突状细胞。我们还将评估纯化的c-di-AMP对树突状细胞活化的影响。 在与万斯实验室（项目3）的合作中，我们将使用ENU诱变来鉴定新的宿主因子 控制着CSP在初步数据中，我们确定Sting（干扰素基因刺激因子）是一种关键的 响应L所必需的主机组件。单核细胞增多症M.结核病和c-di-AMP。在决赛中 目的，我们将测试的假设，即c-di-AMP的生产是必不可少的充分表达L。 单核细胞增多症的发病机制，重要的是，有助于诱导细胞介导的免疫。我们 将检查c-di-AMP作为佐剂的作用。