The role of inositol in Cryptococcus biology and pathogenesis

肌醇在隐球菌生物学和发病机制中的作用

基本信息

批准号：
9903576
负责人：
Chaoyang Xue
金额：
$ 8.72万
依托单位：
RBHS-NEW JERSEY MEDICAL SCHOOL
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：
2016-11-24 至 2021-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9903576
关键词：
Anabolism Animal Model Antifungal Agents Binding Biological Assay Biological Models Biology Biotin Blood Blood - brain barrier anatomy Blood Vessels Brain Cell surface Cells Central Nervous System Infections Cessation of life Complex Cryptococcal Meningitis Cryptococcus Cryptococcus neoformans Development Disease Endothelium Environment Exhibits Extravasation Exudate Fungal Meningitis Future Gene Deletion Genetic Transcription Goals Growth Heart Transplantation Hemostatic function Homeostasis Human Hyaluronic Acid Immune Evasion Immunity Immunocompromised Host In Vitro Infection Inositol Knowledge Label Laboratories Lead Life Ligands Mediating Meningitis Metabolic Pathway Metabolism Mission Modeling Molecular Mus Mutagenesis Mycoses Neuraxis Neurotropism Pathogenesis Pathway interactions Plants Play Polysaccharides Process Production Quantitative Reverse Transcriptase PCR Role Structure System Testing Tight Junctions Transplant Recipients United States National Institutes of Health Virulence Virulence Factors base blood-brain barrier crossing blood-brain barrier penetration capsule chromatin immunoprecipitation deletion library disorder control disorder prevention fungus high throughput screening human pathogen in vivo in vivo imaging system insight monolayer mutant nano-string novel novel strategies pathogen pathogenic fungus promoter transcription factor transcriptome sequencing uptake

项目摘要

ABSTRACT Cryptococcus neoformans is a deadly fungal pathogen that exhibits pronounced neurotropism: it is the most common cause of fungal meningitis, particularly in immunocompromised patients, resulting in over 620,000 deaths annually. How C. neoformans cells traverse the blood-brain barrier (BBB) to infect the central nervous system (CNS) remains poorly understood. Our recent studies implicate inositol – one of the most abundant metabolites in the brain – in this process. In particular, we find that growth of C. neoformans under inositol-rich conditions enhances fungal virulence and that fungal mutants defective in inositol uptake exhibit reduced virulence, reduced capacity to transmigrate from the blood into the brain, and reduced ability to traverse a model BBB in vitro. We also find that C. neoformans compromises tight-junction integrity in vitro, promoting inositol leakage through the brain microvascular endothelial monolayer. Furthermore, we show that inositol induces the expression of fungal cell surface factors involved in virulence, including the polysaccharide capsule, a major fungal virulence factor, and hyaluronic acid (HA), a ligand important for fungal binding to the BBB. Finally, we find that growth on inositol promotes the production of capsule structures involved in immune evasion and that, conversely, C. neoformans mutants defective in inositol uptake elicit enhanced protective immunity during brain infection. Based on these results, we hypothesize that C. neoformans senses and utilizes host inositol to modify the fungal cell surface in a way that promotes penetration of the BBB and development of cryptococcal meningitis. Interestingly, C. neoformans contains an unusually complex inositol uptake system and catabolic pathway, which likely evolved from its utilization of the inositol stores of its plant reservoirs. Thus, this fungus may be uniquely adapted to thrive in the inositol-rich environment of the CNS and to utilize inositol-dependent pathways for pathogenesis. The overarching goal of this proposal is to obtain a detailed understanding of the mechanism by which C. neoformans acquires and utilizes host inositol to establish human brain infection. We propose three Specific Aims: 1) Define inositol sensing and metabolic pathways required for modifying fungal cell surface structure by using a combination of fungal mutagenesis analysis, enzymatic assays, and polysaccharide structural analysis; 2) Characterize the mechanism of inositol promotion in C. neoformans BBB crossing and CNS infection by using an in vitro BBB model system and in vivo animal models; and 3) Define the transcriptional circuits regulating inositol-dependent processes during cryptococcal brain infection by analyzing the expression and localization of fungal inositol factors and identifying transcription factors regulating their expression during infection. Together, these studies will elucidate a novel contribution of a brain metabolite, inositol, to the development of life-threatening fungal meningitis. As such, these studies promise to provide substantial insight into mechanisms by which pathogens cross the BBB and establish CNS infections.

抽象的新型隐球菌是一种致命的真菌病原体，具有明显的向神经性：它是最致命的真菌病原体。真菌性脑膜炎的常见原因，尤其是免疫功能低下的患者，导致超过 620,000 每年死亡人数。新型隐球菌细胞如何穿过血脑屏障（BBB）感染中枢神经系统我们最近的研究表明肌醇是最丰富的物质之一。大脑中的代谢物——在这个过程中，我们特别发现新型隐球菌在富含肌醇的情况下生长。条件增强了真菌毒力，并且肌醇摄取缺陷的真菌突变体表现出降低毒力、从血液转移到大脑的能力降低以及穿过病毒的能力降低我们还发现新型隐球菌会损害体外的紧密连接完整性，从而促进 BBB 体外模型的形成。肌醇通过脑微血管内皮单层渗漏此外，我们还发现肌醇。诱导参与毒力的真菌细胞表面因子的表达，包括多糖胶囊（一种主要的真菌毒力因子）和透明质酸（HA）（一种对真菌与真菌结合很重要的配体） BBB。最后，我们发现肌醇的生长促进了参与免疫的胶囊结构的产生。逃避，相反，肌醇摄取缺陷的新型隐球菌突变体会引发增强的保护作用根据这些结果，我们认为新型隐球菌具有感知能力和免疫力。利用宿主肌醇以促进 BBB 渗透的方式修饰真菌细胞表面，隐球菌脑膜炎的发展提示，新型隐球菌含有异常复杂的肌醇。摄取系统和分解代谢途径，可能是从利用植物的肌醇储备进化而来的因此，这种真菌可能特别适合在中枢神经系统和中枢神经系统富含肌醇的环境中生长。利用肌醇依赖性途径进行发病机制该提案的首要目标是获得一个详细了解新型隐球菌获取和利用宿主肌醇的机制我们提出了三个具体目标：1）定义肌醇传感和代谢。通过使用真菌诱变组合来改变真菌细胞表面结构所需的途径分析、酶分析和多糖结构分析；2) 表征肌醇的作用机制；通过使用体外 BBB 模型系统和在体内促进新型隐球菌 BBB 交叉和 CNS 感染体内动物模型；3) 定义调节肌醇依赖过程的转录回路通过分析真菌肌醇因子的表达和定位来研究隐球菌脑感染这些研究将共同研究在识别感染过程中调节其表达的转录因子。阐明脑代谢物肌醇对危及生命的真菌发展的新贡献因此，这些研究有望为病原体的机制提供深入的见解。穿过血脑屏障并建立中枢神经系统感染。