GENE EXPRESSION AND FUNCTION IN ASPERGILLOSIS

曲霉病中的基因表达和功能

• 批准号: 8893198
• 负责人: Scott G Filler
• 金额: $ 51.31万
• 依托单位: CARNEGIE-MELLON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-15 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8893198
• 关键词: Animal Model; Aspergillosis; Aspergillus fumigatus; Basic Science; Biological; Biological Assay; Biological Process; Biology; Breathing; Candida albicans; Cell surface; Data; Data Set; Defect; Detection; Development; Diagnosis; Diagnostic; Diagnostic radiologic examination; Enzymes; Future; Gene Expression; Gene Expression Profiling; Gene Proteins; Gene Targeting; Genes; Genome; Genomics; Growth; Health; Hematopoietic Stem Cell Transplantation; Immune response; Immunocompromised Host; In Vitro; Infection; Liquid substance; Lung; Modeling; Molds; Molecular Profiling; Morbidity - disease rate; Mus; Mycoses; Organ Transplantation; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Pharmaceutical Preparations; Phenotype; Pilot Projects; Proteins; RNA; Regulatory Pathway; Reproduction spores; Resources; Role; Sampling; Solid; Surface; Testing; Tissues; Toxin; Transcript; Transcription factor genes; Validation; Virulence; Voriconazole; base; chromatin immunoprecipitation; design; enzyme biosynthesis; gene function; immunoregulation; improved; in vitro Assay; in vivo; information model; mortality; mutant; nano-string; pathogen; prospective; research study; response; therapeutic target; therapy development; transcription factor; transcriptome sequencing

DESCRIPTION (provided by applicant): Aspergillus fumigatus is the major invasive mold pathogen of immunosuppressed patients, causing exceptionally high morbidity and mortality. Weakened defenses as a consequence of hematopoietic stem cell transplantation or solid organ transplantation allow growth of inhaled spores in the lung and dissemination. Both therapy and diagnosis are problematic, a situation that contributes to extremely high mortality rates. Our objective is to use A. fumigatus gene expression during infection in a murine model to define virulence genes. We will prioritize genes for functional analysis based on in vivo expression and host response, and then validate the utility of the data through construction of new mutant strains and determination of their virulence potential. These gene products in turn will be high priority targets for therapeutic and diagnostic development. We will use a nanoString nCounter to quantify specific A. fumigatus RNA levels in infected tissue. NanoString estimates of RNA levels are much more sensitive than microarray estimates or current RNA-Seq capability. Our proposed studies focus on three classes of gene products: transcription factors, surface and secreted proteins, and secondary metabolite biosynthetic enzymes. We have chosen transcription factor genes because their products can be connected to target genes and biological functions through expression profiling and chromatin immunoprecipitation. We have chosen surface and secreted protein genes because their products present the most accessible therapeutic targets, and because their possible release into fluids makes them candidate diagnostic targets. We have chosen secondary metabolite genes because their products may have biological activity, such as immunomodulation, that is relevant to pathogenesis. We will develop a reference dataset that explores expression of these genes, and extend the analysis with functional validation through two specific aims. First, we will define A. fumigatus gene expression during lung infection, using a murine model and two sequenced A. fumigatus strains. Second, we will validate functional inferences from expression data through virulence assays of select mutant strains. Our findings will significantly improve the understanding of A. fumigatus infection by providing a set of regulatory pathways, surface/secreted products, and toxins that impact infection. The information will be critical for prioritizing pathways and gene products as targets for therapeutic and diagnostic development, and to connect basic research studies to gene products that are highly relevant to infection.

描述（由申请人提供）：曲曲霉是免疫抑制患者的主要侵入性霉菌病原体，导致发病率异常高。由于造血干细胞移植或固体器官移植而导致的防御能力减弱，从而使肺部吸入孢子的生长并传播。治疗和诊断都是有问题的，这种情况导致了极高的死亡率。我们的目标是在鼠模型中使用感染过程中的烟曲霉基因表达来定义毒力基因。我们将基于体内表达和宿主反应的功能分析确定基因的优先级，然后通过构建新的突变菌株并确定其毒力潜力来验证数据的实用性。这些基因产物反过来将是治疗和诊断发展的高优先级目标。我们将使用纳米弦ncounter来量化受感染组织中特定的烟曲霉RNA水平。 RNA水平的纳米弦估计比微阵列估计值或当前的RNA-Seq能力更敏感。我们提出的研究集中于三类基因产物：转录因子，表面和分泌蛋白，以及继发代谢物生物合成酶。我们选择了转录因子基因，因为它们的产物可以通过表达分析和染色质免疫沉淀连接到靶基因和生物学功能。我们之所以选择表面和分泌的蛋白质基因，是因为它们的产品呈现出最容易获得的治疗靶标，并且由于它们可能释放到液体中，因此它们使它们成为候选诊断靶标。我们之所以选择次生代谢产物基因，是因为它们的产物可能具有与发病机理有关的生物学活性，例如免疫调节。我们将开发一个参考数据集，该数据集探索这些基因的表达，并通过两个特定目的通过功能验证扩展分析。首先，我们将使用鼠模型和两个测序的烟曲霉菌株定义肺部感染过程中的烟曲霉基因表达。其次，我们将通过精选突变菌株的毒力测定来验证表达数据的功能推断。我们的发现将通过提供一组影响感染的调节途径，表面/分泌产物和毒素来显着提高对烟曲霉感染的理解。该信息对于优先考虑途径和基因产品作为治疗和诊断发展的目标至关重要，并将基础研究与与感染高度相关的基因产品联系起来。