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: 10350983
• 负责人: ASIYA GUSA
• 金额: $ 9.84万
• 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/10350983
• 关键词: AIDS/HIV problem; 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; 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; Play; 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 隐球菌属环境真菌，主要在免疫功能低下的人群中引起疾病​​， 其中包括致命的隐球菌性脑膜炎，15% 的艾滋病毒/艾滋病相关死亡都是因吸入而导致的。 进入肺部后，这些真菌必须迅速适应，才能在人类宿主遇到的各种压力下生存， 包括高温应激、pH 值变化和氧化应激。 隐球菌必须逃避宿主免疫防御并抵抗抗真菌药物治疗。 已知在感染期间增强毒力或引起耐药性的隐球菌的变化包括碱基 替换、小插入/缺失和非整倍性转座元件 (TE) 是小的、可移动的 DNA。 大多数真核生物基因组中存在的元件能够引起显着的基因组变化 TE 在隐球菌和其他致病真菌中的潜在作用。 （念珠菌和曲霉）对真菌发病机制或耐药性的影响在很大程度上尚未被探索。 最近发现隐球菌中的 TE 动员是小鼠突变的重要原因 感染模型中报告基因的 TE 突变在高浓度下显着升高。 体外温度（37°宿主相关温度），表明热应激刺激TE在 此外，我们证明 TE 插入是临床耐药性的一个原因。 我们的研究是第一个在体外鉴定 TE 的抗真菌药物雷帕霉素/FK506 和 5-氟胞嘧啶。 动员作为病原真菌感染期间突变的原因此外，TE 诱变。 此前尚未在任何模型酵母物种中描述过对热应激的反应。 在 C. deneoformans 中鉴定出的响应性 TE 包括 DNA 转座子和逆转录转座子，每种都具有 在拟议的研究中，我们寻求不同的动员模式和基因组整合的首选位点。 1) 确定热应激是否是 C. deneoformans 期间 TE 动员增加的主要原因 感染，2) 确定 C. deneoformans 中热应激诱导的 TE 突变的调节因子，以及 3) 确定 TE 是否在体外或宿主感染期间在其他隐球菌物种（新型隐球菌或格特隐球菌）中动员。 阐明增强真菌发病机制或产生耐药性的适应性变化机制是 对于开发和维持有效的抗真菌治疗至关重要，这项研究将进一步加深我们对抗真菌治疗的理解。 隐球菌感染期间出现的压力诱导突变类型可能导致疾病 此外，我们的研究将强调一组患者临床结果的持久性和变化。 可用于识别和定量的实验方法、感染模型和测序工具 多种致病性和非致病性物种的基因突变（TE 和非 TE）。