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.

项目摘要 向量传播疾病占所有感染疾病的17％以上，并造成700,000多人死亡 再过一次。仅蚊子就会导致40万疟疾死亡，并将病毒传播到数百万。 蚊子的媒介能力取决于其生存感染的能力。致病性的破坏性影响 蚊子中肠的入侵是有据可查的15-22，但对蚊子如何容忍知之甚少 压力。肠道干细胞（ISC）介导的中肠上皮修复对于果蝇生存至关重要 病原体的口服摄入33。蚊子中肠上皮包含ISC样细胞27、34、37-39，但其功能性 感染结局的重要性和蚊子生存尚不清楚。我们建议解决这一知识 通过研究对致病性侵袭的蚊子肠道再生反应，媒介生物学的间隙。 关于蚊子中ISC的功能意义的“黑匣子”是基本的一部分 知识差距：物理研究将蚊子中肠视为均匀的整体，而不是 包括多个细胞种群的复杂，区域分区的组织（例如，肠细胞， 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具有 没有被调查。拟议的工作不仅会照亮蚊子上皮的反应 细胞水平，但将为更广泛的媒介生物学界创建新的创新工具。 该项目的前两个目的是（a）表征蚊子下的肠道上皮细胞动力学 体内平衡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。埃及埃及将用作确定哪些刺激的模型（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，哪种遗传途径控制中肠上皮修复以及什么 角色上皮修复在矢量生存和能力中发挥作用。我们的第三个目标是（c）确定特定 功能分化的细胞群对上皮动力学和感染的贡献 回复。我们将使用单细胞RNASEQ/ATACSEQ来确定组成中肠的数量数量 上皮，创建新的转基因线，表达重要细胞类型的荧光标记（肠内分泌） 细胞和ISC），并使用这些线将荧光激活的细胞分类与RNASEQ进行检查以检查 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蚊子再生反应的关键空白 我们的长期目标是确定破坏肠道再生的向量控制策略的新目标 并将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 传播。除了为创新控制目标奠定基础外，我们还将为 更广泛的媒介生物学界，为蚊子中肠生理学中的新发现铺平了道路。