Virulence factor discovery in the secreted proteome of Histoplasma capsulatum

荚膜组织胞浆菌分泌蛋白质组中毒力因子的发现

基本信息

批准号：
8089509
负责人：
Chad A Rappleye
金额：
$ 32.1万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-08-01 至 2013-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8089509
关键词：
Address Affect Alveolar Macrophages Animals Antibiotics Antibodies Biochemical Cells Clinical Data Defense Mechanisms Development Disease Enabling Factors Environment Extracellular Protein Fungal Proteins Gene Deletion Gene Silencing Gene Structure Generations Genes Genetic Genomics Health Histopathology Histoplasma Histoplasma capsulatum Histoplasmosis Human Immune Immunocompetent Immunocompromised Host Incidence Individual Infection Invaded Knowledge Lead Life Lung Lung diseases Mass Spectrum Analysis Measures Methodology Methods Microbe Modeling Modern Medicine Molecular Molecular Genetics Mus Mycoses Oxidative Stress Pathogenesis Performance Phagosomes Phase Production Proteins Proteome Proteomics RNA Interference Regimen Research Proposals Respiratory Tract Infections Reverse Transcriptase Polymerase Chain Reaction Role Science Superoxide Dismutase Systemic disease TYRP1 gene Testing Time Tissues Virulence Virulence Factors Work Yeasts base catalase design extracellular fungus gene function genome sequencing improved in vivo intracellular parasitism killings macrophage mutant novel pathogen prevent protein aminoacid sequence protein expression research study respiratory success tool

项目摘要

DESCRIPTION (provided by applicant): Fungal diseases have emerged as major life-threatening complications of modern medicine as well as respiratory and systemic disease among both immunocompromised and immunocompetent individuals. The dimorphic fungi in particular can cause disease even among otherwise healthy people. Despite this significant impact on human health, surprisingly few fungal gene products have been identified that promote the virulence of medically important fungi. Yeast cells of the dimorphic fungal pathogen, Histoplasma capsulatum, invade, replicate within, and ultimately kill host macrophages. How Histoplasma succeeds in these tasks is unknown due to significant technical limitations which have hindered progress in understanding Histoplasma pathogenesis at a molecular level. We recently developed powerful molecular tools enabling suppression of Histoplasma gene function thereby providing the means to now functionally define the role of genes in virulence. To address this deficiency in our understanding, this research proposal will identify and define additional fungal virulence determinants among the factors secreted by pathogenic Histoplasma yeast. These extracellular factors are prime candidates for affecting the host macrophages and immune defenses. Taking advantage of recently completed genomic sequence information, extracellular factors will be identified through proteomic-based analyses of proteins secreted by Histoplasma yeast. The genes encoding these factors will then serve as targets for gene silencing by RNA interference or gene deletion methodologies to eliminate their production. This ability to create strains of Histoplasma lacking individual proteins permits functional tests to be performed. Strains will be used to infect human and mouse macrophages in culture to assess intracellular parasitism and will be employed in animal infection models of respiratory histoplasmosis. These experiments are designed to assess the contribution of individual factors in Histoplasma pathogenesis. PUBLIC HEALTH RELEVANCE: Our understanding of the mechanisms by which pathogenic fungi infect and cause disease in human hosts is limited which impairs our options to treat life-threatening fungal disease. This proposal will identify molecules released by fungal cells which allow the pathogen to overcome host immune defenses. Improved understanding of the virulence mechanisms employed by pathogenic fungi, specifically the identification of factors that enable them to cause disease, will facilitate the rational design of better antibiotics and regimens to treat clinical fungal infections.

描述（由申请人提供）：真菌疾病已成为现代医学的主要危及生命的并发症，以及免疫功能低下和免疫功能正常个体的呼吸系统疾病和全身疾病。尤其是二态性真菌，即使在其他方面健康的人中也可能引起疾病。尽管对人类健康有重大影响，但令人惊讶的是，很少有真菌基因产物能够增强医学上重要真菌的毒力。二形性真菌病原体荚膜组织胞浆菌的酵母细胞侵入宿主巨噬细胞，在其中复制并最终杀死宿主巨噬细胞。由于显着的技术限制阻碍了在分子水平上理解组织胞浆菌发病机制的进展，组织胞浆菌如何成功完成这些任务尚不清楚。我们最近开发了强大的分子工具，能够抑制组织胞浆菌基因功能，从而提供了现在在功能上定义基因在毒力中的作用的方法。为了解决我们理解中的这一缺陷，本研究计划将在致病性组织胞浆菌分泌的因子中识别和定义其他真菌毒力决定因素。这些细胞外因子是影响宿主巨噬细胞和免疫防御的主要候选因子。利用最近完成的基因组序列信息，将通过对组织胞浆菌酵母分泌的蛋白质进行基于蛋白质组学的分析来鉴定细胞外因子。然后，编码这些因子的基因将作为基因沉默的靶标，通过 RNA 干扰或基因删除方法来消除它们的产生。这种产生缺乏单个蛋白质的组织胞浆菌菌株的能力允许进行功能测试。菌株将用于感染培养中的人类和小鼠巨噬细胞，以评估细胞内寄生，并将用于呼吸道组织胞浆菌病的动物感染模型。这些实验旨在评估个体因素在组织胞浆菌发病机制中的贡献。公共卫生相关性：我们对病原真菌感染人类宿主并引起疾病的机制的了解有限，这损害了我们治疗危及生命的真菌疾病的选择。该提案将鉴定真菌细胞释放的分子，这些分子使病原体能够克服宿主的免疫防御。更好地了解病原真菌的毒力机制，特别是识别导致疾病的因素，将有助于合理设计更好的抗生素和治疗临床真菌感染的方案。