The role of stem-cell mediated midgut repair in the dynamics of mosquito infections

干细胞介导的中肠修复在蚊子感染动态中的作用

• 批准号: 10404118
• 负责人: Nicolas Buchon
• 金额: $ 51.04万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-05 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10404118
• 关键词: ATAC-seq; Acute; Address; Aedes; Affect; Age; Animal Model; Animals; Antimitotic Agents; Area; Bacteria; Biology; Blood; Cell Differentiation process; Cells; Cessation of life; Communicable Diseases; Communities; Competence; Complex; Culicidae; Disease; Drosophila genus; Enterocytes; Enteroendocrine Cell; Epithelial; Epithelial Cells; Epithelial Physiology; Equilibrium; Fluorescence-Activated Cell Sorting; Genetic; Genetic Transcription; Goals; Homeostasis; Human; Infection; Intestines; Investigation; Knowledge; Longevity; Malaria; Mediating; Midgut; Modeling; Natural regeneration; Oral; Oral Ingestion; Outcome; Pathogenicity; Pathway interactions; Phenotype; Physiological; Physiology; Play; Population; Regenerative response; Reporting; Research; Resolution; Role; Stimulus; Stress; Testing; Tissues; Transgenic Organisms; Vector-transmitted infectious disease; Vectorial capacity; Virus; Work; cell type; epithelial repair; feeding; gastrointestinal epithelium; human pathogen; innovation; microbial; mosquito-borne; novel; oral infection; oral pathogen; pathogen; progenitor; repaired; response; single-cell RNA sequencing; stem cell proliferation; stem cells; stem-like cell; success; tool; transcriptome sequencing; transmission process; vector; vector control; vector mosquito; virtual

Project Summary Vector-borne diseases account for more than 17% of all infectious disease and cause more than 700,000 deaths annually 1. Mosquitoes alone cause 400,000 malaria deaths and transmit viruses to hundreds of millions 2. The vectorial capacity of mosquitoes depends on their ability to survive infection. The damaging effects of pathogenic invasion of the mosquito midgut are well-documented 15-22, but little is known about how mosquitoes tolerate this stress. Intestinal stem cell (ISC) mediated midgut epithelial repair is essential for Drosophila survival following oral ingestion of pathogens 33. The mosquito midgut epithelium contains ISC-like cells 27, 34, 37-39, but their functional significance for infection outcomes and mosquito survival is unknown. We propose to address this knowledge gap in vector biology by investigating the mosquito gut regenerative response to pathogenic invasion. The “black box” regarding the functional significance of ISCs in the mosquito midgut is part of a fundamental knowledge gap: physiological studies treat the mosquito midgut as a homogeneous whole, rather than a complex, regionally compartmentalized tissue comprised of multiple cell populations (e.g. enterocytes, enteroendocrine cells, and ISCs). The specific contributions of these cell types to gut-pathogen interactions have not been investigated. The proposed work will not only illuminate mosquito epithelial responses to infection at the cellular level but will lead to the creation of new and innovative tools for the broader vector biology community. The first two aims of this project are (A) to characterize gut epithelial cell dynamics in mosquitoes under conditions of homeostasis and oral infection and (B) to evaluate the role of midgut epithelial repair in mosquito infection outcomes. Aedes aegypti will be used as a model to determine what stimuli (including human pathogens) affect gut epithelial turnover rates, whether post-infection repair rebuilds the gut homeostatically or alters epithelial composition, what genetic pathways control midgut epithelial repair, and what role epithelial repair plays in vector survival and competence. Our third aim is (C) to determine the specific contributions of functionally differentiated cell populations to epithelial dynamics and infection response. We will use single-cell RNAseq/ATACseq to establish how many cell types compose the midgut epithelium, create new transgenic lines expressing fluorescent markers for important cell types (enteroendocrine cells and ISCs), and, using these lines, couple fluorescence-activated cell sorting with RNAseq to examine the transcriptional response of the three major cell types (enterocytes, enteroendocrine cells, and ISCs) to infection. Our study will fill a critical gap in our understanding of mosquito midgut regenerative responses to pathogenic invasion. Our long-term goal is to identify new targets for vector control strategies that disrupt gut regeneration and reduce survival of infected mosquitoes below the critical incubation threshold required for pathogen transmission. In addition to laying the groundwork for innovative control targets, we will create tools for the broader vector biology community, paving the way for novel discoveries in mosquito midgut physiology.

项目概要 媒介传播疾病占所有传染病的 17% 以上，导致超过 70 万人死亡 每年 1. 仅蚊子就导致 40 万人死于疟疾，并将病毒传播给数亿人 2. 蚊子的传播能力取决于它们在病原体感染的破坏性影响下的生存能力。 蚊子中肠的入侵已有充分记录 15-22，但人们对蚊子如何忍受这种情况知之甚少 肠干细胞（ISC）介导的中肠上皮修复对于果蝇的存活至关重要。 口服摄入病原体 33. 蚊子中肠上皮含有 ISC 样细胞 27, 34, 37-39，但它们的功能 对于感染结果和蚊子生存的意义尚不清楚，我们建议解决这一问题。 通过研究蚊子肠道对病原体入侵的再生反应来弥补媒介生物学的空白。 关于蚊子中肠中 ISC 功能意义的“黑匣子”是基本原理的一部分 知识差距：生理学研究将蚊子中肠视为一个均匀的整体，而不是一个 由多个细胞群（例如肠上皮细胞、 肠内分泌细胞和 ISC）。这些细胞类型对肠道病原体相互作用的具体贡献有 拟议的工作不仅会阐明蚊子上皮对感染的反应。 细胞水平，但将为更广泛的载体生物学界创造新的创新工具。 该项目的前两个目标是 (A) 表征蚊子肠道上皮细胞动态 稳态和口腔感染的条件，以及（B）评估中肠上皮修复的作用 蚊子感染结果将被用作模型来确定哪些刺激（包括 人类病原体）影响肠道上皮周转率，感染后修复是否重建肠道 稳态或改变上皮组成，什么遗传途径控制中肠上皮修复，以及什么 我们的第三个目标是（C）确定上皮修复在载体存活和能力中的具体作用。 功能分化的细胞群对上皮动力学和感染的贡献 我们将使用单细胞 RNAseq/ATACseq 来确定中肠有多少种细胞类型。 上皮细胞，创建表达重要细胞类型（肠内分泌细胞）荧光标记的新转基因系 细胞和 ISC），并且使用这些细胞系，将荧光激活细胞分选与 RNAseq 结合起来，以检查 三种主要细胞类型（肠细胞、肠内分泌细胞和 ISC）对感染的转录反应。 我们的研究将填补我们对蚊子中肠对病原体的再生反应的理解的关键空白 我们的长期目标是确定破坏肠道再生的媒介控制策略的新目标。 并将受感染蚊子的存活率降低到病原体所需的临界孵化阈值以下 除了为创新控制目标奠定基础外，我们还将创建工具。 更广泛的媒介生物学界，为蚊子中肠生理学的新发现铺平了道路。