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

• 批准号: 7902025
• 负责人: Chad A Rappleye
• 金额: $ 32.52万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 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/7902025
• 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; 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; Race; 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; public health relevance; research study; respiratory; success; tool

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的个体的呼吸道和全身性疾病。特别是双态真菌，即使在其他健康的人中也会引起疾病。尽管这种对人类健康的重大影响，令人惊讶的是，很少有真菌基因产物已被确定为促进医学上重要的真菌的毒力。酵母细胞的双态真菌病原体，荚膜组织胞浆菌，入侵，复制内，并最终杀死宿主巨噬细胞。组织胞浆菌如何在这些任务中取得成功是未知的，这是由于重大的技术限制，阻碍了在分子水平上理解组织胞浆菌发病机制的进展。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，从而提供了现在功能上定义基因在毒力中的作用的手段。为了解决我们理解中的这一缺陷，本研究提案将在致病性组织胞浆菌酵母分泌的因子中鉴定和定义其他真菌毒力决定因子。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者。利用最近完成的基因组序列信息，细胞外因子将被确定通过蛋白质组学为基础的分析蛋白质分泌的组织胞浆菌酵母。然后，编码这些因子的基因将通过RNA干扰或基因缺失方法作为基因沉默的靶标，以消除它们的产生。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物中的人和小鼠巨噬细胞，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。这些实验旨在评估组织胞浆菌发病机制中个体因素的作用。公共卫生相关性：我们对致病真菌感染人类宿主并导致疾病的机制的理解是有限的，这削弱了我们治疗危及生命的真菌疾病的选择。这项提案将确定真菌细胞释放的分子，这些分子允许病原体克服宿主的免疫防御。提高对致病真菌毒力机制的认识，特别是确定致病因素，将有助于合理设计更好的抗生素和治疗方案，以治疗临床真菌感染。