RAS GENE FUNCTION IN FUNGAL DIMORPHISM

该提案的长期目标是了解分子 cAMP调节二形细胞分化的机制， 病原真菌，荚膜组织胞浆菌。H. capsulatum是使役 病原体的疾病，组织胞浆菌病，最常见的原发性 西半球的系统性真菌感染数百万人 世界各地的人都感染了这种病菌， 报告的艾滋病患者播散性组织胞浆菌病病例中， 以惊人的速度增长。土壤中的微生物以 多细胞丝状菌丝体，但发现的唯一形式的真菌 是单细胞的芽殖酵母从菌丝体阶段开始 阁下于荚膜是非致病性的并且只有酵母相是寄生的， 控制相变的因素对于理解 致病性 H.在实验室中，通过控制 培养温度：酵母菌在37 ℃生长，菌丝体在25 ℃生长 摄氏度通过改变孵化温度， 一个相位到另一个相位被感应。最有希望的候选人之一， 对温度做出反应并触发 形态发生，是cAMP。酵母培养物暴露于引起 细胞内cAMP水平，诱导转化为菌丝体在 37摄氏度的非允许温度。其具体目的是 建议是确定RAS功能是否涉及细胞内 H.荚膜 在酵母中，RAS蛋白控制GTP刺激的腺苷酸环化酶活性 并且至少一个功能性RAS基因对于细胞活力是必需的， 增殖和孢子萌发。为了确定ras功能是否 参与H.在荚膜中，RAS基因将被 通过标准分子生物学技术克隆和表征。 这些基因在形态发生过程中的表达将通过北方 分析和生化方法将用于确定是否RAS H. capsulatum在cAMP效应子途径中起作用。

The long-term objective of this proposal is to understand the molecular mechanism by which cAMP modulates differentiation of the dimorphic, pathogenic fungus, Histoplasma capsulatum. H. capsulatum is the causative agent of the disease, histoplasmosis, one of the most common primary systemic fungal infections in the Western Hemisphere. Millions of people throughout the world have become infected with the organism, and the number of reported cases of disseminated histoplasmosis in patients with AIDS is increasing at an alarming rate. The organism exists in soil as multicellular, filamentous mycelia, but the only form of the fungus found in infected tissue is unicellular, budding yeast. Since the mycelial phase of H. capsulatum is nonpathogenic and only the yeast phase is parasitic, factors which control the phase transition are critical for understanding pathogenicity. H. capsulatum is easily maintained in the laboratory by controlling the temperature of incubation; yeast grow at 37 degrees C and mycelia at 25 degrees C. By shifting the temperature of incubation, the transition from one phase to the other is induced. One of the most promising candidates for the transducing signal that responds to temperature and triggers morphogenesis, is cAMP. Exposure of yeast cultures to agents which raise the intracellular level of cAMP, induces the transition to mycelia at the nonpermissive temperature of 37 degrees C. The specific aim of this proposal is to determine whether ras function is involved in the cellular response to cAMP in H. capsulatum. In yeast, RAS proteins control GTP-stimulated adenylate cyclase activity and at least one functional RAS gene is essential for cell viability, proliferation and spore germination. To determine whether ras function is involved in the differentiation of H. capsulatum, the RAS genes will be cloned and characterized by standard molecular biological techniques. Expression of these genes during morphogenesis will be assessed by Northern analysis, and biochemical methods will be utilized to determine whether RAS genes in H. capsulatum function in the cAMP effector pathway.