Exploring the Structure and Function of Parasitic Nematode Exosomes

淋巴丝虫病(LF)是一种由蚊子传播的被忽视的热带疾病，由丝虫引起，包括 马来丝虫；全世界有超过1.2亿人感染，在70个地方病流行中，有超过14亿人面临风险 国家。目前使用大规模药物管理的控制策略已经降低了许多 但低频仍然是一个重大的全球卫生问题。人们认识到需要新的战略来 控制由寄生线虫引起的LF等疾病。这个项目的重点是寄主寄生虫 寄生虫感染期间的界面，LF的发展和持续，代表了一个令人兴奋的新方向 调查这一领域。我们提出了一种新的机制，通过它来调节宿主免疫。 系统，通过一种特定类型的胞外囊泡递送的小的、调节的RNA和蛋白质 外显体。我们的初步数据表明，感染阶段的马来丝虫幼虫分泌外切体， 这些外体含有不同的miRNA和蛋白质，而不同的寄生虫miRNA是潜在的靶标 寄主基因。此外，这些外切体被宿主巨噬细胞内化，并引发一种特定的调节 表型。这项提案的总体目标是定义寄生虫外周体通过 哺乳动物内丝虫的生命周期，并探索其生物活性的机制基础。 在特定的目标1中，我们建议对分泌的外切体的小RNA和蛋白质货物进行分析 利用小RNA深度测序(RNA-Seq)和序列分析相结合的方法研究哺乳动物内马来丝虫的生活史 蛋白质组学分析以确定寄生虫外切体传递的宿主操纵的分子介体。在……里面 具体目标2，我们将确定外切体对宿主免疫细胞介质的生物活性机制 回应。我们将检测宿主巨噬细胞对寄生虫外切体的摄取，以揭示这些小泡是如何 被宿主细胞内化，然后通过使用基因敲击来利用小鼠模型的遗传能力 OUTS对外切体在宿主巨噬细胞中诱导的调制表型的机械性洞察 内部化。 该项目的长期影响将是对马来芽胞杆菌生物学的新知识和新的 可以在新的LF控制策略中利用的分子。此外，我们在这里描述的机制可能 在动物、人类和植物寄生线虫中保守，可用于广谱 控制应用程序。

Lymphatic filariasis (LF) is a mosquito-borne Neglected Tropical Disease caused by filarial worms including Brugia malayi; over 120 million people worldwide are infected, with over 1.4 billion at risk in 70 endemic countries. Current control strategies employing mass drug administration have reduced prevalence in many areas, but LF remains a significant global health concern. There is a recognized need for new strategies to control LF and other diseases caused by parasitic nematodes. This project focuses on the host-parasite interface during parasite infection, development and persistence of LF and represents an exciting new direction for investigating this field. We propose a novel mechanism by which B. malayi modulates the host immune system, through small, regulatory RNAs and proteins delivered via a specific type of extracellular vesicle called exosomes. Our preliminary data demonstrate that infective stage larvae of B. malayi secrete exosomes, that these exosomes contain a diverse miRNA and protein cargo and that distinct parasite miRNA potentially target host genes. Further, these exosomes are internalized by host macrophages and elicit a specific modulatory phenotype. The overall goals of this proposal are to define the cargo of parasite exosomes secreted across the intra-mammalian Brugia life cycle and probe the mechanistic basis for their bioactivity. In Specific Aim 1, we propose to profile the small RNA and protein cargo of secreted exosomes across the intra-mammalian B. malayi life cycle using a combination of small RNA deep-sequencing (RNA-Seq) and proteomic profiling to identify the molecular mediators of host manipulation delivered by parasite exosomes. In Specific Aim 2, we will define the mechanisms of exosome bioactivity on cellular mediators of the host immune response. We will examine uptake of parasite exosomes by host macrophages to reveal how these vesicles are internalized by host cells, then leverage the genetic capacity of the murine model by using genetic knock outs to generate mechanistic insight into the modulatory phenotype elicited in host macrophages by exosome internalization. The long-term impact of the project will be new knowledge of B. malayi biology and the exposure of new molecules that may be exploited in novel LF control strategies. Further, the mechanisms we describe here may be conserved across animal, human and plant parasitic nematodes and could be utilized for broad-spectrum control applications.