Identifying Wolbachia effectors that facilitate host infection

识别促进宿主感染的沃尔巴克氏体效应子

• 批准号: 10613456
• 负责人: Irene Newton
• 金额: $ 38.53万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-10 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10613456
• 关键词: Actins; Aedes; Alphaproteobacteria; Animals; Arbovirus Infections; Bacteria; Bioinformatics; Biological Assay; Biology; Cell Line; Cell physiology; Cells; Cellular biology; Collaborations; Communicable Diseases; Coupling; Culicidae; Data; Dengue Virus; Disease Vectors; Drosophila genus; Drosophila melanogaster; Environment; Evolution; Future; Gene Expression; Genetic; Genome; Goals; Growth; Immunoprecipitation; In Vitro; Infection; Insecta; Invaded; Investigation; Mass Spectrum Analysis; Mediating; Modeling; Modification; Molecular; Oocytes; Parasites; Persons; Phenotype; Planet Earth; Planets; Protein Secretion; Proteins; Proteomics; RNA Viruses; Regulation; Reproduction; Research; Specificity; Symbiosis; System; Type IV Secretion System Pathway; Vaccines; Viral; Virus; Virus Replication; Wolbachia; Work; candidate identification; comparative genomics; differential expression; disorder control; fly; genetic manipulation; host colonization; human disease; infancy; interest; mortality; mosquito-borne pathogen; nanomachine; next generation; non-Native; overexpression; pathogen; prevent; programs; symbiont; transgene expression; transmission process; vector competence; vector control

PROJECT SUMMARY/ABSTRACT Wolbachia pipientis is an obligate intracellular alpha-proteobacterium that infects 40-60% of insect species on the planet. Wolbachia infection inhibits RNA virus replication in insects, a phenomenon known as pathogen blocking. Therefore, Wolbachia infected mosquitos are being released in many parts of the world to control the spread of human diseases. Importantly, although the mechanism behind Wolbachia’s virus inhibition is not known, Wolbachia must colonize the host and be efficiently maternally transmitted in order for pathogen blocking to work. Our long-term goals are to identify the mechanisms used by Wolbachia to establish infection. To that end, we focus on the type IV secretion system (T4SS), a molecular nanomachine used by Wolbachia to inject proteins, termed effectors, into the host cellular environment. Via these secreted effectors, the host cell is modified, allowing Wolbachia to invade and persist. Our previous work identified and characterized the first secreted effector in Wolbachia (WalE1) and established an important correlation between T4SS gene expression and expression of secreted effectors. Wolbachia effectors and the T4SS are upregulated during host pupation. Strikingly, overexpression of WalE1 in Wolbachia-infected flies facilitated colonization of the developing oocyte and maternal transmission. These results led to our central hypothesis that Wolbachia uses secreted effectors to establish and maintain host colonization. Towards this hypothesis, we have identified Wolbachia effectors across the genus, and used a large-scale growth screen to further identify Wolbachia effectors. We have discovered that many Wolbachia effectors are clade specific, but many are shared between the insect associated strains. In collaboration with Peter Christie (UTH), we developed a heterologous secretion assay for Wolbachia effectors, allowing us to confirm interactions between the type IV coupling protein VirD4 and candidate Wolbachia effectors. Guided by strong preliminary data, we propose to pursue three Specific Aims to identify and characterize Wolbachia effectors, and their host targets, using the power of the Drosophila system. We will (1) determine the Wolbachia secretome, (2) predict effectors and their conservation across Wolbachia infecting insects, and (3) identify host targets and effectors that facilitate infection by Wolbachia. Studies of Wolbachia - host interactions are still in their infancy despite the recognized contributions of endosymbiotic associations to insect reproduction and evolution, and the ability to alter vector competence. These proposed studies will significantly advance our understanding of how Wolbachia employs its T4SS to establish infection, a necessary prerequisite to pathogen blocking.

项目摘要/摘要 细管沃尔巴克氏杆菌是一种专性胞内α-蛋白细菌，可感染40%-60%的昆虫。 这颗星球。沃尔巴克氏菌感染抑制昆虫体内的RNA病毒复制，这种现象被称为病原体 阻挡。因此，受沃尔巴克氏菌感染的蚊子正在世界许多地区被释放，以控制 人类疾病的传播。重要的是，尽管沃尔巴克氏菌病毒抑制背后的机制并不是 众所周知，沃尔巴克氏菌必须定植在宿主上，并通过母体有效传播，才能使病原体 挡着去上班了。我们的长期目标是确定沃尔巴克氏菌用来确定感染的机制。 为此，我们将重点放在IV型分泌系统(T4SS)上，这是沃尔巴克氏杆菌用来 将被称为效应器的蛋白质注入宿主细胞环境。通过这些分泌的效应器，宿主细胞 经过改良，允许沃尔巴克氏菌入侵并持续存在。 我们以前的工作鉴定并鉴定了沃尔巴克氏菌中的第一个分泌型效应器(Wale1)，并建立了 T4SS基因表达与分泌型效应物的表达之间存在重要的相关性。沃尔巴克氏菌 效应器和T4SS在寄主化蛹过程中上调。令人惊讶的是，WalE1在 感染沃尔巴克氏菌的苍蝇促进了发育中的卵母细胞的定居和母体传播。这些 结果导致了我们的中心假设，沃尔巴克氏杆菌使用分泌的效应器来建立和维持宿主 殖民主义。对于这一假设，我们已经确定了整个属的沃尔巴克氏菌效应器，并使用了 大规模生长屏进一步鉴定沃尔巴克氏菌效应器。我们发现许多沃尔巴克氏菌 效应器是支系特有的，但许多是昆虫相关菌株共享的。与 彼得·克里斯蒂(UTH)，我们开发了一种沃尔巴克氏菌效应器的异源分泌试验，使我们能够 确认IV型偶联蛋白VirD4与候选沃尔巴克氏菌效应器之间的相互作用。指导原则 强大的初步数据，我们建议追求三个具体目标来识别和表征沃尔巴克氏菌 效应器和它们的宿主目标，使用果蝇系统的力量。我们将(1)确定 沃尔巴克氏菌分泌组，(2)预测感染沃尔巴克氏菌的昆虫中的效应物及其保护，以及(3) 确定促进沃尔巴克氏菌感染的宿主靶标和效应物。 沃尔巴克氏杆菌与宿主相互作用的研究仍处于初级阶段，尽管已有公认的贡献 内共生与昆虫繁殖和进化的关系，以及改变媒介能力的能力。 这些拟议的研究将大大促进我们对沃尔巴克氏杆菌如何利用其T4SS来 建立感染，这是病原体阻断的必要前提。