Wolbachia disrupts eukaryotic endolysosomal membrane dynamics

沃尔巴克氏菌破坏真核细胞内溶酶体膜动力学

基本信息

批准号：
10667824
负责人：
Vincent Joseph Starai
金额：
$ 18.42万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-01-15 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10667824
关键词：
Address Adult Antibiotics Bacteria Bacterial Proteins Binding Biochemical Biological Models Biology Biotinylation Brugia Brugia malayi Cellular biology Cessation of life Complex Coupled Data Development Disease Dissection Elephantiasis Endosomes Ensure Eukaryota Family Filarial Elephantiases Filariasis Genes Genetic Goals Human In Vitro Individual Intracellular Membranes Ivermectin Knowledge Laboratories Lipid Biochemistry Membrane Membrane Fusion Membrane Proteins Microbiology Modeling Molecular Nematoda Ocular Onchocerciasis Organism Outcome Pathogenicity Pathway interactions Physiology Population Process Proteins Publishing Reagent Regulation Reporting Reproduction Resistance Saccharomycetales Symbiosis System Technology Testing Tissues Vacuole Wolbachia Work Yeasts endosymbiont genetic manipulation global health human disease human pathogen in vivo inhibitor insight novel pathogen protein degradation protein function protein protein interaction protein transport receptor tool trafficking yeast protein

项目摘要

PROJECT SUMMARY Filarial nematodes of the family Onchocercidae cause debilitating human diseases, such as lymphatic filariasis. As approximately 150 million individuals are currently infected with these nematodes, obtaining in- depth knowledge of pathogen biology will serve to address a global health issue. It is known that the filarial nematode, Brugia malayi, harbors an intracellular endosymbiotic bacterium of the Wolbachia genus, and this relationship is essential; clearance of Wolbachia from the nematode with antibiotics leads to eventual nematode death. Understanding the mechanisms by which Wolbachia maintains its intracellular survival within nematodes would therefore likely provide an important avenue towards controlling pathogenic nematode populations, but both Brugia and Wolbachia are not amenable to genetic manipulations. Discoveries of important bacterium:nematode interactions at the molecular level, therefore, have proven exceedingly difficult. In this proposal, our goals are to utilize proteins from Wolbachia to genetically and biochemically dissect conserved pathways of endolysosomal membrane dynamics in yeast. These secreted “effector” proteins are known to alter host processes in order to support the survival of the bacterium in the eukaryotic host and to ensure its own reproduction, and are therefore potent reagents that impact eukaryotic physiology. To this end, my laboratory has employed the budding yeast, Saccharomoyces cerevisiae (Sce), as a model system towards the discovery of bacterial proteins that modulate eukaryotic cellular biology, with a focus on those proteins which inhibit intracellular membrane fusion and protein trafficking pathways. In a previous screen of candidate wBm secreted effector proteins, we have already identified proteins from wBm that have the ability to manipulate eukaryotic biology. In this work, we show that one such protein, wBm0152, strongly inhibits endosome:vacuole trafficking pathways in vitro. This inhibition appears to result from modulation of the conserved ESCRT complex. As wBm is known to alter membrane dynamics in its host during its symbiosis, and coupled with the fact that regulation of membrane dynamics is strongly conserved throughout eukaryotes, the detailed genetic, molecular, and biochemical studies carried out in this proposal will be applicable to wBm:B. malayi interactions, and thus, human filarial diseases. Finally, leveraging our laboratory's strengths in microbiology, cellular biology, and protein/ lipid biochemistry, we will carefully detail the biochemical activity of this Wolbachia-derived ESCRT modulator and identify important regulators and binding partners in yeast, which are likely conserved in Brugia. This work will begin to describe heretofore unknown wBm:B. malayi interactions, thus providing novel insight into not only Brugia physiology, but also provide new insight into ESCRT-dependent activities in eukaryotes.

项目概要盘尾丝虫科的丝虫线虫会导致人类衰弱疾病，例如淋巴疾病由于目前大约有 1.5 亿人感染了这些线虫，因此，众所周知，对病原体生物学的深入了解将有助于解决全球健康问题。线虫，马来丝虫，含有沃尔巴克氏菌属的细胞内内共生细菌，并且这种细菌用抗生素从线虫中清除沃尔巴克氏体会导致最终的线虫。了解沃尔巴克氏体在线虫中维持细胞内存活的机制。因此可能为控制致病线虫种群提供重要途径，但是布鲁氏菌和沃尔巴克氏菌都不适合进行重要的基因操作。因此，细菌与线虫之间的相互作用已被证明很难超过分子水平。在这项提案中，我们的目标是利用沃尔巴克氏体的蛋白质进行遗传和生化解剖酵母中溶酶体膜动力学的保守途径是这些分泌的“效应”蛋白。已知可以改变宿主过程以支持细菌在真核宿主中的生存并确保其自身的繁殖，因此是影响真核生理学的有效试剂。我的实验室采用了芽殖酵母，酿酒酵母（Sce），作为模型系统调节真核细胞生物学的细菌蛋白质的发现，重点是那些能够调节真核细胞生物学的蛋白质抑制细胞内膜融合和蛋白质运输途径。在之前对候选 wBm 分泌效应蛋白的筛选中，我们已经鉴定了蛋白质在这项工作中，我们展示了一种这样的蛋白质， wBm0152，在体外强烈抑制内体：液泡运输途径。众所周知，wBm 可以改变宿主体内的膜动力学。它的共生关系，再加上膜动力学的调节在整个过程中都高度保守的事实真核生物，本提案中进行的详细遗传、分子和生化研究将适用最后，利用我们实验室的优势。在微生物学、细胞生物学和蛋白质/脂质生物化学中，我们将仔细详细介绍这种源自沃尔巴克氏体的 ESCRT 调节剂并鉴定了酵母中的重要调节剂和结合伴侣，可能在 Brugia 中保守。这项工作将开始描述迄今为止未知的 wBm:B。因此，不仅为布鲁氏菌生理学提供了新的见解，而且还为ESCRT依赖性提供了新的见解真核生物中的活动。