Mitchondrial targeting by the vacuolating cytotoxin of Helicobacter Pylori

幽门螺杆菌空泡细胞毒素的线粒体靶向

基本信息

批准号：
9263698
负责人：
Robin L. Holland
金额：
$ 3.8万
依托单位：
UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-04-01 至 2020-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9263698
关键词：
Address Bacteria Bacterial Toxins Binding Biochemical Carrier Proteins Cell physiology Cell surface Cells Cellular biology Complex Cytosol Cytotoxin Development Disease Doctor of Philosophy Endosomes Epithelial Cells Exotoxins Experimental Designs Fractionation Gene Deletion Genes Health Helicobacter pylori Homeostasis Human Hybrids Illinois Infection Innate Immune Response Integration Host Factors Intoxication Knock-out Knowledge Label Laboratories Life Magnetism Mammalian Cell Mediating Membrane Metabolic Control Mitochondria Mitochondrial Proteins Modeling Nuclear Pathogenesis Pathogenicity Pathway interactions Physiologic pulse Physiological Protein Import Proteins Proteome Proteomics RNA Interference Reporting Research Risk Stomach Surface System Targeted Toxins Testing Time Toxin Transport Vesicles Universities Vesicle Vesicle Transport Pathway Viral Virulence Factors base cell injury clinically relevant ferrite ferromagnetism insight interest knock-down malignant stomach neoplasm mitochondrial dysfunction nanoparticle novel strategies novel therapeutics pathogen pathogenic bacteria predictive modeling protein transport public health relevance therapy development trafficking uptake

项目摘要

DESCRIPTION (provided by applicant): This application proposes studies to identify the mechanism by which the Helicobacter pylori vacuolating cytotoxin (VacA) is trafficked to mitochondria in host epithelial cells, where the toxin induces mitochondrial damage. VacA binds to the surface of host cells and is internalized into the endolysosomal system, but a major gap in knowledge to be addressed in this proposal is the mechanism by which the toxin targets and localizes to mitochondria. We and others have failed to detect VacA within the host cell cytosol, suggesting that this membrane-interacting, pore-forming toxin is not taken up into mitochondria using existing pathways used by endogenous proteins that are imported from the cytosol. Rather, our current model predicts that VacA is transported from the cell surface to mitochondria by vesicular trafficking. Given that vesicular trafficking pathways from the cell-surface to mitochondria have not previously been identified, this application proposes studies to evaluate the hypothesis that VacA-containing vesicles (VCVs) are dynamically re-modeled from early- endosomal-like compartments to trafficking vesicles that are competent for targeting mitochondria. Working in the laboratory of Dr. Steven Blanke at the University of Illinois at Urbana-Champaign, I will test the prediction of this hypothesis using a set of studies proposed within 2 Specific Aims: 1) In Aim 1, studies are proposed to test the prediction that, if VCVs are dynamically remodeled, the proteome of VCVs will change as a function of time to become enriched with proteins that mediate mitochondrial targeting and/or vesicular trafficking. In support of the proposed studies, I have developed and optimized new approaches for magnetically isolating VCVs enriched in VacA labeled with ferromagnetic nano-particles. 2) In Aim 2, I will test the prediction that if VCVs become progressively enriched with cellular proteins required for mitochondrial targeting, then knockdown or knockout of these cellular proteins by RNA interference or gene deletion will result in decreased or blocked VacA localization to mitochondria, and, VacA-mediated disruption of mitochondrial function. Completion of the proposed studies will address a major gap in our understanding of cellular intoxication by VacA, which is a major virulence factor of H. pylori, a pathogen of significant health concern given human infection increases the risk for the development of gastric cancer. These studies will also reveal, potentially for the first time, an intracellular trafficking mechanism by which proteins move from the surface of host cells to the mitochondria, which may be relevant to the broader class of mitochondrial-acting, pore-forming toxins generated by clinically relevant pathogenic bacteria. Identifying the mechanisms and importance of bacterial toxin-mediated modulation of host cells is an important step towards developing therapies for blocking toxin activities that contribute to pathogenesis.

描述（通过应用程序证明）：该应用提出的是确定幽门螺杆菌的幽门螺杆菌在宿主上皮细胞中的线粒体的机制，毒素是毒素会诱导线粒体损害的其他人未能检测到宿主细胞细胞溶胶中的VACA，这表明使用内源性的途径，我们目前的模型从细胞表面传输了，使用了从Thetos进口的内源性的途径。通过囊泡运输的线粒体，从细胞表面到线粒体的途径，含Vaca的囊泡（VCV）是动态重新建模的在博士的实验室中，是使用2个特定目的中提出的一组研究的假设为假设：1）在AIM 1中，提出了研究以测试以下预测：如果VCV进行了动态重塑，VCV的蛋白质组将变化尽可能多。富含线粒体靶向和/或囊泡运输的富含维特蛋白，我已经开发了并优化了磁性分离的新方法，从而富含铁磁性的VCV逐渐富集了VCV。线粒体靶向的必需品，然后通过RNA干扰或封闭对线粒体的VACA定位，敲除这些地窖蛋白，VACA介导的破坏了线粒体功能的主要差距。幽门螺杆菌的感染增加了胃癌发展的风险。相关的致病细菌。