Intracellular Proliferation of the fungal pathogen Histoplasma capsulatum

真菌病原体荚膜组织胞浆菌的细胞内增殖

• 批准号: 10583569
• 负责人: Chad A Rappleye
• 金额: $ 37.45万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-10 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10583569
• 关键词: Address; Amino Acid Permease; Amino Acid Transporter; Amino Acids; Attenuated; Carbon; Catabolism; Cells; Citric Acid Cycle; Coccidioidomycosis; Complement; Consumption; Data; Diagnosis; Disease; Environment; Enzymes; Funding Opportunities; Genes; Genetic; Genetic Screening; Genetic Transcription; Genus Hippocampus; Gluconeogenesis; Glutamate Dehydrogenase; Glutamates; Glycolysis; Goals; Growth; Histoplasma; Histoplasma capsulatum; Histoplasmosis; Immune system; Immunocompetent; Immunocompromised Host; Impairment; In Vitro; Individual; Infection; Innate Immune System; Invaded; Investigation; Isotope Labeling; Lung; Macrophage; Measurement; Metabolic; Metabolic Pathway; Metabolism; Mitochondria; Molecular; Molecular Biology; Molecular Genetics; Mutation; Mycoses; Nutrient; Nutritional; Nutritional Requirements; Pathogenesis; Pathway interactions; Peptides; Phagocytes; Phagosomes; Proliferating; RNA Interference; Reaction; Resources; Respiration; Respiratory Disease; Saccharomyces cerevisiae; Source; Substrate Specificity; Systemic disease; Testing; Therapeutic; Time; Virulence; Yeasts; alpha ketoglutarate; desert fever; fungus; genetic testing; in vivo; interdisciplinary approach; metabolic profile; metabolomics; mouse model; mutant; new therapeutic target; novel strategies; novel therapeutics; parasitism; pathogen; pathogenic fungus; permissiveness; prevent; residence; response; transcriptomics; uptake

PROJECT SUMMARY Histoplasma capsulatum is a fungal pathogen that infects both immunocompromised and immunocompetent individuals. The innate immune system alone is ineffective in controlling Histoplasma yeasts because Histoplasma invades and thrives within host phagocytic cells. Histoplasma’s parasitism of macrophages requires acquisition and metabolism of suitable nutritional resources from the host cell, yet the host molecules that serve as nutrients for are largely unknown. Preliminary investigations have shown intracellular yeasts are gluconeogenic and likely consume amino acids, particularly compounds that are related to glutamate. These findings provide a critical foothold for identifying the host molecules consumed by Histoplasma yeasts. This proposal takes a multidisciplinary approach, integrating evidence from molecular biology, genetics, and metabolomics to determine the host molecules available to Histoplasma yeasts within the phagosome and how they are metabolized to meet the carbon and energy needs within host cells. The intracellular growth requirement for glutamate dehydrogenase activity narrows the potential host molecules for metabolic carbon to glutamate-related amino acids or glutamate-generating host molecules. Transcriptional and metabolite profiling will be combined with functional tests to define the metabolic pathways necessary for Histoplasma proliferation within host macrophages. Isotopic labeling of host metabolites will be used to follow their import and subsequent incorporation into Histoplasma metabolism intermediates over time to demonstrate the flux of carbon from macrophage to intracellular yeasts and identify host substrate entry points into yeast central carbon metabolism. These findings will be genetically tested by characterization of Histoplasma’s amino acid and peptide transporters and RNAi-based interference with candidate host metabolite import into intracellular Histoplasma yeast cells. Together these data will provide multiple lines of evidence to define how Histoplasma yeasts exploit the phagosome as a replication-permissive intracellular niche during host infection. This proposal is submitted in response to Funding Opportunity Announcement (FOA) PA-19-083 “Novel approaches to understand, prevent, treat, and diagnose coccidioidomycosis (Valley Fever) and other select endemic fungal infections.” The proposal answers the announcement’s goal to address the pathogenesis of endemic fungi with the ultimate goal of advancing the field towards solutions for treatment of endemic mycoses. Of the dimorphic endemic fungal pathogens, Histoplasma is the best molecularly characterized fungus and it has the most advanced molecular genetics to facilitate functional testing for mechanistic studies. The results of our studies will reveal new disease treatment avenues by highlighting transport and metabolic reactions essential for the narrow metabolism imposed on Histoplasma by its residence within macrophage phagosomes. Our genetic tests have shown that impairment of these critical metabolic reactions reduces Histoplasma proliferation in the mammalian lung, demonstrating the therapeutic potential.

项目摘要 荚膜组织胞浆菌是一种真菌病原体，感染免疫功能低下和免疫活性 个体单独的先天免疫系统在控制组织胞浆菌方面是无效的， 组织胞浆菌在宿主吞噬细胞内侵入并繁殖。组织胞浆菌对巨噬细胞的寄生 需要从宿主细胞获得和代谢合适的营养资源，然而宿主分子 作为营养物质的生物大部分是未知的。初步研究表明，细胞内酵母是 可能会消耗氨基酸，特别是与谷氨酸有关的化合物。这些 这一发现为鉴定组织胞浆菌消耗的宿主分子提供了关键的立足点。 这项建议采取了多学科的方法，整合了分子生物学，遗传学， 代谢组学，以确定宿主分子可用于组织胞浆菌酵母内的吞噬体，以及如何 它们被代谢以满足宿主细胞内的碳和能量需求。细胞内生长 对谷氨酸脱氢酶活性的需要使代谢碳的潜在宿主分子变窄， 谷氨酸相关氨基酸或谷氨酸生成宿主分子。转录和代谢产物 分析将与功能测试相结合，以确定组织胞浆菌所需的代谢途径。 在宿主巨噬细胞内增殖。宿主代谢物的同位素标记将用于跟踪其输入 并随后随着时间的推移掺入组织胞浆菌代谢中间体中，以证明 从巨噬细胞到细胞内酵母的碳，并确定宿主底物进入酵母中心的入口点 碳代谢这些发现将通过组织胞浆菌的氨基酸特征进行遗传学检验。 和肽转运蛋白和基于RNAi的干扰候选宿主代谢物进入细胞内 组织胞浆菌酵母细胞。这些数据将提供多条证据来确定组织胞浆菌 酵母在宿主感染期间利用吞噬体作为允许复制的细胞内小生境。 本提案是为了响应资助机会公告（FOA）PA-19-083“小说”而提交的。 方法来了解，预防，治疗和诊断球孢子菌病（山谷热）和其他选择 地方性真菌感染”该提案回答了公告的目标，即解决 地方性真菌的最终目标是推进该领域的解决方案，以治疗地方性 真菌病组织胞浆菌是二型地方性真菌病原体中分子特征最好的 真菌，它具有最先进的分子遗传学，以促进机制研究的功能测试。 我们的研究结果将揭示新的疾病治疗途径，通过强调运输和代谢 组织胞浆菌驻留在巨噬细胞内，对组织胞浆菌的狭义代谢至关重要 吞噬体我们的基因测试表明，这些关键的代谢反应的损害， 组织胞浆菌在哺乳动物肺中的增殖，证明了治疗潜力。