Stress-induced transposon mobilization in the human fungal pathogen Cryptococcus

人类真菌病原体隐球菌中应激诱导的转座子动员

• 批准号: 10590596
• 负责人: ASIYA GUSA
• 金额: $ 9.88万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-11 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10590596
• 关键词: Aneuploidy; Antifungal Agents; Area; Aspergillus; Biological Assay; Brain; Candida; Cessation of life; Chemicals; Clinical; Cryptococcal Meningitis; Cryptococcus; Cryptococcus gattii; Cryptococcus neoformans; DNA; DNA Insertion Elements; DNA Methylation; DNA Sequence Alteration; DNA Transposable Elements; DNA Transposons; Disease; Drug resistance; Elements; Environment; Evolution; FK506; Flucytosine; Frequencies; Fungal Drug Resistance; Gene Expression; Genetic Variation; Genome; Genomic DNA; Genomics; Goals; Growth; HIV/AIDS; Heat Stress Disorders; Heat-Shock Response; High temperature of physical object; Human; Immune; Immune system; Immunocompromised Host; In Vitro; Induced Mutation; Infection; Inhalation; Invertebrates; Life; Lung; Maps; Mediating; Meningitis; Modeling; Molecular Chaperones; Movement; Mus; Mutagenesis; Mutation; Mycoses; Organ; Organism; Oxidative Stress; Pathogenesis; Pathogenicity; Patient-Focused Outcomes; Patients; Pharmacotherapy; Phenotype; Population; RNA Interference; Recurrence; Relapse; Reporter; Reporter Genes; Reporting; Research; Retrotransposon; Role; Serial Passage; Sirolimus; Site; Stress; System; Temperature; Variant; Virulence; Waxes; Yeast Model System; Yeasts; base; causal variant; drug isolation; environmental stressor; fungus; gene function; genetic approach; genome sequencing; genome-wide; human disease; in vivo; insertion/deletion mutation; mortality; mouse model; mutant; next generation; novel; pathogenic fungus; response; tool; trait; transposon/insertion element; whole genome

Project Abstract Cryptococcus species are environmental fungi that cause disease primarily in immunocompromised populations, including a deadly cryptococcal meningitis that contributes to 15% of HIV/AIDS-related deaths. When inhaled into the lungs, these fungi must adapt rapidly to survive a variety of stresses encountered in the human host, including high temperature stress, changes in pH and oxidative stress. In cases of persistent disease, Cryptococcus must evade host immune defenses and resist antifungal drug treatment. Adaptive genomic changes in Cryptococcus known to enhance virulence or cause drug resistance during infection include base substitutions, small insertions/deletions and aneuploidy. Transposable elements (TEs) are small, mobile DNA elements present in the genomes of most eukaryotic organisms that are capable of causing significant genomic changes and phenotypic variation. The potential role of TEs in Cryptococcus and other pathogenic fungal species (Candida and Aspergillus) in contributing to fungal pathogenesis or drug resistance is largely unexplored. We recently identified TE mobilization in Cryptococcus deneoformans as a significant cause of mutation in a murine model of infection. Mutations by TEs in reporter genes for drug resistance were dramatically elevated at high temperature (37° host-relevant temperature) in vitro, suggesting that heat stress stimulates TE mobility in the cryptococcal genome. Additionally, we demonstrated TE insertion as a cause of drug resistance to clinical antifungal agents rapamycin/FK506 and 5-fluorocytosine in vitro. Our study was the first to identify TE mobilization as a cause of mutation during infection in a pathogenic fungus. In addition, TE mutagenesis in response to heat stress had not been described previously in any model yeast species. Remarkably, the heat- responsive TEs identified in C. deneoformans include both DNA transposons and retrotransposons, each with distinct modes of mobilization and preferred sites of genomic integration. In the proposed research, we seek to 1) determine whether heat stress is the primary cause of increased TE mobilization during C. deneoformans infection, 2) identify regulators of heat stress-induced TE mutagenesis in C. deneoformans, and 3) determine whether TEs mobilize in other cryptococcal species (C. neoformans or C. gattii) in vitro or during host infection. Elucidating the mechanisms of adaptive change that enhance fungal pathogenesis or enable drug resistance is critical in developing and maintaining effective antifungal treatments. This study will further our understanding of the types of stress-induced mutations that arise during cryptococcal infection that may contribute to disease persistence and variations in clinical outcomes for patients. In addition, our study will highlight a set of experimental approaches, infection models and sequencing tools that can be used to identify and quantitate genetic mutations (TE and non-TE) in a broad range of pathogenic and non-pathogenic species.

项目摘要 隐球菌是环境真菌，在免疫功能低下的人群中引起疾病​​的主要真菌， 包括致命的隐球菌脑膜炎，造成与HIV/艾滋病相关的15％的死亡。继承时 在肺部，这些真菌必须迅速适应以在人类宿主中遇到的各种压力中生存， 包括高温应激，pH变化和氧化应激。在持续性疾病的情况下 隐球菌必须逃避宿主的免疫输液并抵抗抗真菌药物治疗。自适应基因组 在感染过程中已知可以增强病毒或引起耐药性的隐球菌的变化包括基础 取代，小插入/缺失和非整倍性。转座元素（TES）小，移动DNA 大多数真核生物的基因组中存在的元素，能够引起重要的基因组 变化和表型变化。 TES在加密环球和其他致病真菌物种中的潜在作用 （念珠菌和曲曲霉）在导致真菌发病机理或耐药性的贡献很大程度上是出乎意料的。我们 最近确定了deneoformans中的TE动员为鼠突变的重要原因 感染模型。在耐药性中，TES在耐药性中通过TES突变在高中显着升高 体外温度（37°宿主温度），表明热应力刺激了TE迁移率 隐球菌基因组。此外，我们证明了TE插入是对临床耐药性的原因 抗真菌剂雷帕霉素/FK506和5-氟环胞素体外。我们的研究是第一个识别 动员作为致病真菌感染过程中突变的原因。另外，诱变 值得注意的是，先前在任何模型的酵母菌中都没有描述过热压力。 在脱氧杆菌中鉴定出的反应性TE都包括DNA转座子和逆转录座子，每个TE都有 动员的不同模式和基因组整合的首选位点。在拟议的研究中，我们试图 1）确定热应激是否是脱氧杆菌中TE动员增加的主要原因 感染，2）鉴定脱氧杆菌中热应激诱导的TE诱变的调节剂，3）确定 无论是在体外还是在宿主感染期间动员其他隐球菌（Neoformans或C. gattii）中动员。 阐明增强真菌发病机理或启用耐药性的自适应变化机制是 对于开发和维持有效的抗真菌治疗至关重要。这项研究将进一步了解 在隐球菌感染期间出现的压力诱导的突变的类型，可能导致疾病 患者临床结局的持久性和变化。此外，我们的研究将突出一组 实验方法，感染模型和测序工具，可用于识别和定量 在广泛的致病性和非致病物种中，基因突变（TE和非TE）。