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）：烟曲霉是免疫抑制患者的主要侵袭性霉菌病原体，导致极高的发病率和死亡率。造血干细胞移植或实体器官移植导致防御能力减弱，导致吸入孢子在肺部生长和传播。治疗和诊断都存在问题，这种情况导致极高的死亡率。我们的目标是利用小鼠模型感染期间的烟曲霉基因表达来定义毒力基因。我们将根据体内表达和宿主反应对基因进行功能分析，然后通过构建新的突变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成为治疗和诊断开发的高度优先目标。我们将使用 nanoString nCounter 来量化受感染组织中特定的烟曲霉 RNA 水平。 NanoString 对 RNA 水平的估计比微阵列估计或当前的 RNA-Seq 功能灵敏得多。我们提出的研究重点关注三类基因产物：转录因子、表面和分泌蛋白以及次级代谢产物生物合成酶。我们选择转录因子基因是因为它们的产物可以通过表达谱和染色质免疫沉淀与靶基因和生物功能联系起来。我们选择了表面和分泌蛋白基因，因为它们的产物提供了最容易接近的治疗靶点，并且因为它们可能释放到液体中使它们成为候选诊断靶点。我们选择次级代谢基因是因为它们的产物可能具有与发病机制相关的生物活性，例如免疫调节。我们将开发一个参考数据集来探索这些基因的表达，并通过两个特定目标扩展分析和功能验证。首先，我们将使用小鼠模型和两个已测序的烟曲霉菌株来定义肺部感染期间烟曲霉的基因表达。其次，我们将通过选定突变株的毒力测定来验证表达数据的功能推论。我们的研究结果将通过提供一系列影响感染的调控途径、表面/分泌产物和毒素，显着提高对烟曲霉感染的理解。这些信息对于优先考虑途径和基因产物作为治疗和诊断开发的目标，以及将基础研究与与感染高度相关的基因产物联系起来至关重要。