Synthetic Genetic Analysis of the RAM network in C. albicans

白色念珠菌 RAM 网络的合成遗传分析

• 批准号: 7754756
• 负责人: ANUJ KUMAR
• 金额: $ 19.28万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-18 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7754756
• 关键词: AIDS/HIV problem; Achievement; Affect; Alleles; Antifungal Therapy; Binding; Biological Assay; Candida albicans; Candidate Disease Gene; Candidiasis; Characteristics; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Data; Development; Diploidy; Disease; Drug resistance; Epithelium; Esophageal; Filament; Genes; Genetic; Genetic Phenomena; Genetic Screening; Goals; HIV; Haploidy; Heterozygote; Human; Hyphae; Immunocompromised Host; Infection; Life; Methods; Morbidity - disease rate; Morphogenesis; Morphology; Mutagenesis; Mutation; Nature; Organism; Oropharyngeal; Pain; Pathogenesis; Pathway interactions; Patients; Penetration; Phenotype; Play; Process; Protein Kinase; Proteins; Recurrence; Regulation; Regulatory Pathway; Relative (related person); Research; Role; Saccharomyces cerevisiae; Screening procedure; Shapes; Signal Pathway; Staging; Testing; Virulence; Yeast Model System; Yeasts; base; chemical genetics; chromatin immunoprecipitation; design; esophagus ulcer; gel mobility shift assay; genetic analysis; improved; insight; interest; mutant; novel; novel strategies; null mutation; oral lesion; pathogen; promoter; public health relevance; response; transcription factor; AIDS/HIV problem; Achievement; Affect; Alleles; Antifungal Therapy; Binding; Biological Assay; Candida albicans; Candidate Disease Gene; Candidiasis; Characteristics; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Data; Development; Diploidy; Disease; Drug resistance; Epithelium; Esophageal; Filament; Genes; Genetic; Genetic Phenomena; Genetic Screening; Goals; HIV; Haploidy; Heterozygote; Human; Hyphae; Immunocompromised Host; Infection; Life; Methods; Morbidity - disease rate; Morphogenesis; Morphology; Mutagenesis; Mutation; Nature; Organism; Oropharyngeal; Pain; Pathogenesis; Pathway interactions; Patients; Penetration; Phenotype; Play; Process; Protein Kinase; Proteins; Recurrence; Regulation; Regulatory Pathway; Relative (related person); Research; Role; Saccharomyces cerevisiae; Screening procedure; Shapes; Signal Pathway; Staging; Testing; Virulence; Yeast Model System; Yeasts; base; chemical genetics; chromatin immunoprecipitation; design; esophagus ulcer; gel mobility shift assay; genetic analysis; improved; insight; interest; mutant; novel; novel strategies; null mutation; oral lesion; pathogen; promoter; public health relevance; response; transcription factor

DESCRIPTION (provided by applicant): Candida albicans is an important opportunistic fungal pathogen that causes both mucosal and invasive disease in immunocompromised patients. Nearly all HIV/AIDS patients will develop oropharyngeal (OPC) or esophageal candidiasis during their lifetime. Because of its recurrent nature, the development of drug-resistant OPC/EC is an important problem, leading to current efforts toward developing novel approaches to therapy based on modulation of C. albicans virulence. For such strategies to be successful, an improved understanding of C. albicans pathogenesis is needed. The ability of C. albicans to undergo morphogenesis from yeast to hyphal form is clearly required for the organism to cause disease. However, the regulatory mechanisms that control hypha formation are incompletely defined. In this project, we have developed a novel large scale genetic screening strategy to identify interactions between pairs of genes that affect hyphal development. The strategy is based on a genetic phenomenon called complex haploinsufficiency (CHI) which occurs when heterozygous mutations in two separate genes cause a more severe phenotype than either single mutation by itself. We have used transposon mutagenesis to generate a large set of double mutants starting with a strain lacking one allele of CBK1, a protein kinase involved in hyphal development. Preliminary data indicate that this screen has identified a number of novel genes involved in hyphal development and has provided evidence supporting the hypothesis that CBK1 acts in parallel with the protein kinase A pathway to regulated hyphal development. In this application, we propose to further develop this screen by pursuing the following specific aims: 1) Identify genes that display complex haploinsufficient interactions with CBK1 in C. albicans; 2) Test the hypothesis that a CHI screen of CBK1 will identify targets of the Cbk1-dependant transcription factor Ace2; and 3) Test the hypothesis that the RAM (CBK1) and cAMP-PKA signaling pathways function in parallel during hypha development. PUBLIC HEALTH RELEVANCE: Candida albicans is an important opportunistic fungal pathogen that causes disease in immunocompromised patients. Nearly all HIV/AIDS patients will develop oropharyngeal or esophageal candidal infection during their lifetime. The ability of C. albicans to cause disease is directly related to its ability to change its shape from a round yeast form to an elongated filamentous form. The proposed research will probe the genetic mechanisms by which C. albicans undergoes this change. We have designed a novel genetic screening strategy to identify genes involved in filament formation that would not otherwise be apparent from standard screening approaches. The mechanistic information generated by this study should help design new antifungal therapies targeted at processes involved in pathogenesis.

描述（由申请人提供）：白色念珠菌是一种重要的机会真菌病原体，可在免疫功能低下的患者中引起粘膜和侵入性疾病。几乎所有艾滋病毒/艾滋病患者都会在其一生中发展出口咽（OPC）或食管念珠菌病。由于其反复发作的性质，耐药OPC/EC的发展是一个重要的问题，这是一个重要的问题，导致目前基于白色念珠菌毒力的调节，为开发新的治疗方法而努力。为了使这些策略成功，需要对白色念珠菌发病机理有了改进的了解。显然，白色念珠菌从酵母到菌丝形式进行形态发生的能力显然需要生物引起疾病。但是，控制菌丝形成的调节机制未完全定义。在这个项目中，我们开发了一种新型的大规模遗传筛查策略，以识别影响菌丝发育的基因对之间的相互作用。该策略是基于一种称为复杂单倍不足（CHI）的遗传现象，该现象发生在两个单独基因的杂合突变引起比单个突变本身更严重的表型时发生的。我们已经使用了转座子诱变来产生大量的双重突变体，从缺乏CBK1等位基因的菌株开始，这是一种参与菌丝发育的蛋白激酶。初步数据表明，该屏幕已经确定了许多参与菌丝发育的新基因，并提供了支持CBK1与蛋白激酶并行作用的证据。在此应用程序中，我们建议通过追求以下特定目的进一步开发此屏幕：1）确定在白色念珠菌中与CBK1表现出复杂单倍弹性相互作用的基因； 2）检验了CBK1的卡屏将识别CBK1依赖性转录因子ACE2的靶标的假设； 3）检验以下假设：RAM（CBK1）和CAMP-PKA信号通路在菌丝发育过程中并行起作用。公共卫生相关性：白色念珠菌是一种重要的机会真菌病原体，可在免疫功能低下的患者中引起疾病​​。几乎所有艾滋病毒/艾滋病患者都会在其一生中发展出口咽或食管念珠菌感染。白色念珠菌引起疾病的能力与其从圆形酵母形式变为细长丝状形式的能力直接相关。拟议的研究将探究白色念珠菌经历这种变化的遗传机制。我们设计了一种新型的遗传筛查策略，以识别与细丝形成有关的基因，从标准筛选方法中看不明会明显。这项研究产生的机械信息应有助于设计针对发病机理过程的新的抗真菌疗法。