DIMORPHISM AND VIRULENCE IN HISTOPLASMA CAPSULATUM

荚膜组织胞浆菌的二态性和毒力

• 批准号: 3144482
• 负责人: GEORGE S KOBAYASHI
• 金额: $ 17.49万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-05-01 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3144482
• 关键词: cell transformation; environmental adaptation; fungal genetics; gene expression; genetic manipulation; genetic strain; histoplasmosis; laboratory mouse; microorganism genetics; microorganism sexual dimorphism; molecular cloning; protein biosynthesis; proteins; stress proteins; virulence

Histoplasma capsulatum, a dimorphic fungus, causes disease in humans that has a wide range of clinical manifestations. The long-term goal of this application is to develop an approach towards improving the outcome of histoplasmosis using biochemical and molecular techniques based on our earlier demonstration that virulent infection requires a phase transition at 37 degrees C in the host. We want to elucidate how genes coding for stress proteins and transition-specific (ts) proteins contribute to the morphologic transition and to the different degrees of pathogenicity in isolates from patients and soil. These studies will be of value in development of drugs that interfere with establishment of infection. There are three specific aims: 1. STudy genes involved in the early stages of the phase transition: a. Thermal adaptation - heat shock genes (hsg): clone and characterize the heat shock 83 gene and determine how regulation of its expression and maturation during phase transition at various temperatures in different isolates is related to thermotolerance and pathogenicity. Determine the physiological role the induction of hs proteins play in the different degrees of thermotolerance, morphogenesis, mitochondrial membrane integrity and virulence. b. Phase transition - clone and characterize ts gene that are expressed in the first 6 hrs of the mycelium to yeast differentiation process and determine the pattern of their expression in strains, including the transitional-defective PCMS strain, that show different levels of thermotolerance and pathogenicity. Within this group we expect to find genes that contain heat shock elements in their regulatory regions. 2. Determine the effect of disruption of genes involved in the phase transition and in virulence by: a. Establishing a transformation protocol - design a procedure that will yield an efficient transformation system in order to analyze the role in vitro-mutated genes play in morphogenesis and pathogenicity. b. In vitro mutagenesis to determine its effect on phase transition and adaption - analyze the effect mutagenesis has on the biochemical events that take place after the temperature shift. 3. Determine the role that hs and ts genes play in virulence by: a. Evaluating the in vitro-mutated strains in mice.

荚膜组织胞浆菌是一种二型真菌，可引起人类疾病， 临床表现广泛。 长期目标是 应用是为了制定一种方法，以改善结果， 组织胞浆菌病的生物化学和分子技术的基础上，我们 先前的证明表明，致命的感染需要一个相变， 在37摄氏度的主机。 我们想阐明基因如何编码 应激蛋白和过渡特异性（TS）蛋白有助于 形态转变和不同程度的致病性， 从病人和土壤中分离出来。 这些研究将在以下方面具有价值： 开发干扰感染建立的药物。 有三个具体目标： 1. 研究参与相变早期阶段的基因： a.热适应-热休克基因（hsg）的克隆与鉴定 热休克83基因，并确定如何调节其表达， 在不同温度下的相变过程中的成熟 菌株的耐热性和致病性有关。 确定 hs蛋白的诱导在不同的 耐热性、形态发生、线粒体膜完整性的程度 和毒性。 B.相变-克隆并表征在大肠杆菌中表达的ts基因， 菌丝体向酵母分化过程的前6小时， 确定它们在菌株中的表达模式，包括 过渡缺陷型PCMS菌株，其显示不同水平的 耐热性和致病性。 在这个群体中，我们希望找到 在其调控区含有热休克元件的基因。 2.确定参与阶段的基因的破坏的影响 转化和毒力： a.建立一个转换协议-设计一个程序， 产生一个有效的转换系统，以分析在 体外突变的基因在形态发生和致病性中起作用。 B.体外诱变以确定其对相变的影响， 适应-分析诱变对生化事件的影响 发生在温度变化之后。 3.通过以下方式确定hs和ts基因在毒力中的作用： a.评估小鼠体外突变株。