Genetic basis and pathogenicity of invasive growth in yeast

描述(申请人提供)：真菌是临床感染的主要来源，特别是在免疫系统受损的患者中。真菌用来定植人类宿主的机制尚不完全清楚，但被认为经常涉及侵入性生长。具体地说，许多真菌能够附着和穿透表面，例如人体组织的表面。酿酒酵母是了解真菌侵袭性生长的环境诱因和遗传机制的有价值的模型，它被认为是在免疫低下的人类中定植的。虽然酿酒酵母参比菌株S288C没有表现出侵入性生长，但我们发现许多生态和遗传多样性的菌株可以侵入性生长，其特征的表达通常取决于特定的环境条件。在这项建议中，(目标1)我们使用基因作图和基因工程的组合来识别导致酿酒酵母临床分离株侵袭性生长变异的基因。我们在一个由2,880个分离物组成的小组中进行了我们的遗传图谱研究，这些分离物来自于所有10个可能的成对组合中的5个不同菌株的交配。一旦确定了因果基因座，我们将使用基因工程技术将这些基因座解析为特定的基因和遗传变异。(目的2)然后，我们通过将野生分离株和工程菌株感染到动物模型中来测试已识别的因果变异的潜在相关性。我们将把这些菌株接种到大量蜡蛾幼虫中，这些幼虫通常用于研究微生物的毒力，并测量这些菌株导致疾病或死亡的程度。拟议研究的完成将提供有关侵入性生长的遗传和环境原因的详细信息，并将阐明已确定的等位基因的潜在临床相关性。相关性：真菌致病病例正在增加，到目前为止，在人类感染中发现了500多种真菌。遗传学方法为鉴定真菌致病的分子机制提供了强有力的工具。然而，常见的机会性病原体，如白色念珠菌，由于无法在实验室中有性繁殖而受到遗传系统的重大限制。酿酒酵母的分离株在它们表现出的致病性转变的类型以及这些特征表达的条件方面表现出很大的变异性。为了确定与真菌附着和穿透表面有关的遗传和环境因素，这被认为有助于发病，我们将使用从免疫受损的人类身上采集的多个酿酒酵母分离株进行基因图谱实验。这项拟议的研究将提高对真菌发病机制的总体理解，并可能确定新的抗真菌药物的潜在靶点。

DESCRIPTION (provided by applicant): Fungi are a major source of clinical infections, especially among patients with compromised immune systems. The mechanisms fungi use to colonize human hosts are not fully understood, but are thought to often involve invasive growth. Specifically, many fungi are capable of attaching to and penetrating surfaces, such as those of human tissues. Saccharomyces cerevisiae, which is known to colonize immunocompromised humans, is a valuable model for understanding the environmental triggers and genetic mechanisms that underlie invasive growth in fungi. Although the S. cerevisiae reference strain S288C does not exhibit invasive growth, we have found that many ecologically and genetically diverse isolates can grow invasively, with expression of the trait often dependent on specific environmental conditions. In this proposal, (Aim 1) we use a combination of genetic mapping and genetic engineering to identify genes that cause variability in invasive growth among clinical isolates of S. cerevisiae. We conduct our genetic mapping studies in a panel of 2,880 segregants derived from the mating of 5 diverse strains in all 10 possible pairwise combinations. Once causal loci have been identified, we will use genetic engineering techniques to resolve these loci to specific genes and genetic variants. (Aim 2) We then test the potential relevance of the identified causal variants by infecting wild isolates and engineered strains into an animal model. We will inoculate the strains into a large number of wax moth larvae, which are commonly used to study the virulence of microbes, and measure the extent to which the strains cause sickness or death. Completion of the proposed research will provide detailed information about the genetic and environmental causes of invasive growth, and will also shed light on the potential clinical relevance of the identified alleles. Relevance: Cases of fungal pathogenesis are on the rise, with more than 500 species of fungi identified in human infections to date. Genetic approaches provide powerful tools for identifying the molecular mechanisms underlying pathogenesis in fungi. However, common opportunistic pathogens, such as Candida albicans, suffer from major limitations as genetic systems due to their inability to sexually reproduce in th lab. Isolates of S. cerevisiae exhibit substantial variability in the types of pathogenicity traitsthey exhibit, as well as the conditions in which these traits are expressed. To determine genetic and environmental factors that are involved in fungi attaching to and penetrating surfaces, which is thought to contribute to pathogenesis, we will perform genetic mapping experiments using multiple isolates of S. cerevisiae that were sampled from immunocompromised humans. The proposed research will improve general understanding of fungal pathogenesis and may identify potential targets for new antifungal drugs.