Comparative and functional genomics of C. neoformans

描述（由申请人提供）：真菌病原体新型隐球菌在艾滋病患者中引起危及生命的感染的高发病率，因此这种真菌对全世界超过4000万感染艾滋病毒的人构成重大威胁。此外，近缘种C.格特氏菌最近已成为免疫功能正常人群的主要病原体。我们的长期目标是获得知识，这将导致新的战略，以打击真菌感染。特别是，我们希望详细了解哺乳动物宿主中病原体生长所需的因素，以确定阻断感染的关键目标。在这方面，铁的供应是宿主环境的一个重要指标，也是病原体的一种必需营养素。因此，我们计划研究C. neoformans，作为了解铁在真菌发病机制中的作用的模型。铁水平对于C.这是因为可利用性影响多糖荚膜的大小，而多糖荚膜是真菌的主要毒力因子。我们的假设是，铁对毒力和生长的影响是通过一个由主铁反应调节器Cir1（隐球菌铁调节器）控制的网络发生的。我们相信，Cir1和这个网络的特征将确定铁的收购机制，是必不可少的C。导致疾病。我们的第一个具体目标是描述Cir1的关键功能。我们将具体确定Cir1是否直接结合铁，以及该蛋白质是否结合编码铁摄取功能的靶基因的启动子区域。这项工作将包括蛋白质组学方法，以确定与Cir1相互作用的调控因子。这些因子可能参与调控铁获得的靶基因和与毒力相关的其他功能。我们的第二个具体目标将表征的Hap转录因子复合物在铁收购血红素的作用。HAP基因的表达受Cir1调控，我们有初步数据表明这些基因是血红素使用所必需的。我们的第三个具体目标将采用转录分析，以检查Cir1在使用铁源（血红素和转铁蛋白），是丰富的主机中的作用。最后，我们将构建突变体，是在基因编码的候选功能，在感染过程中使用血红素和转铁蛋白缺陷。可能的目标（例如，血红素加氧酶），且我们预期我们在AIM 3中微阵列工作将鉴定另外的功能。在这项工作的结论，我们将有一个更深入的了解的调控网络，连接铁的毒力，我们将知道的功能，C。在感染过程中从宿主体内偷取铁。这些信息将突出铁相关的关键功能，可以有针对性地治疗隐球菌病。公共卫生相关性：该项目的相关性来自于控制免疫系统受损的人类真菌感染的迫切需要。特别是，4000万或更多感染艾滋病毒的人很有可能死于真菌疾病。该研究将专门研究通过靶向真菌病原体在感染期间获得营养铁的能力来控制感染的潜力。

DESCRIPTION (provided by applicant): The fungal pathogen Cryptococcus neoformans causes a high incidence of life-threatening infections in AIDS patients and this fungus therefore poses a major threat to the > 40 million people worldwide who are infected with HIV. In addition, the related species C. gattii has recently emerged as a primary pathogen of immunocompetent people. Our long-term goal is to acquire knowledge that will lead to new strategies to combat fungal infections. In particular, we want to develop a detailed understanding of the factors required for growth of pathogens in mammalian hosts in order to identify key targets to block infection. In this regard, iron availability is an important indicator of the host environment as well as an essential nutrient for pathogens. Therefore, we plan to investigate iron regulation and acquisition in C. neoformans, as a model for understanding the role of iron in fungal pathogenesis. Iron levels are particularly important for the pathogenesis of C. neoformans because availability influences the size of the polysaccharide capsule that is the major virulence factor of the fungus. Our hypothesis is that the influence of iron on virulence and growth occurs through a network controlled by the master iron-responsive regulator Cir1 (Cryptococcus iron regulator). We believe that the characterization of Cir1 and this network will identify mechanisms of iron acquisition that are essential for C. neoformans to cause disease. Our first specific aim is to characterize the key functions of Cir1. We will specifically determine whether Cir1 directly binds iron and whether the protein binds the promoter regions of target genes encoding iron uptake functions. This work will include proteomic approaches to identify regulatory factors that interact with Cir1. These factors may participate in the regulation of target genes for iron acquisition and other functions relevant to virulence. Our second specific aim will characterize the role of the Hap transcription factor complex in iron acquisition from heme. The expression of the HAP genes is regulated by Cir1 and we have preliminary data that these genes are required for heme use. Our third specific aim will employ transcriptional profiling to examine the role of Cir1 in the use of iron sources (heme and transferrin) that are abundant in the host. Finally, we will construct mutants that are defective in genes encoding candidate functions for the use of heme and transferrin during infection. Likely targets (e.g., heme oxygenase) have been identified and we anticipate that our microarray work in aim 3 will identify additional functions. At the conclusion of this work, we will have a deeper understanding of the regulatory network that links iron to virulence, and we will know the functions that C. neoformans uses to steal iron from the host during infection. This information will highlight the key iron-related functions that can be targeted to treat Cryptococcal disease. PUBLIC HEALTH RELEVANCE: The relevance of this project comes from the pressing need to control fungal infections in humans with impaired immune systems. In particular, the 40 million or more people infected with HIV have a high chance of succumbing to fungal disease. The research will specifically examine the potential to control infections by targeting the ability of fungal pathogens to acquire the nutrient iron during infection.