The opportunist Candida albicans: yeast proliferation and nutritional signaling

机会主义白色念珠菌：酵母增殖和营养信号

• 批准号: 9188792
• 负责人: JULIA R KOEHLER
• 金额: $ 44.25万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-12-01 至 2018-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9188792
• 关键词: Affect; Affinity; Affinity Chromatography; Alleles; Anti-Retroviral Agents; Antifungal Agents; Area; Attention; Biochemical; Biological; Biological Testing; Candida; Candida albicans; Candidiasis; Cell Cycle Progression; Cell division; Cells; Co-Immunoprecipitations; Collection; DNA biosynthesis; Data; Disease; Drops; Elements; Epitopes; Esophageal; Esophagus; Eukaryota; Filament; Frequencies; Genes; Genetic Epistasis; Genetic Transcription; Goals; Growth; HIV; Highly Active Antiretroviral Therapy; Homologous Gene; Human; Hyphae; Immunocompromised Host; Lateral; Location; Malignant Neoplasms; Mass Spectrum Analysis; Mediator of activation protein; Morphogenesis; Nutritional; Opportunistic Infections; Organelles; Oropharyngeal; Pain; Pathway interactions; Patients; Pharyngeal structure; Phosphotransferases; Population; Process; Production; Proliferating; Proteins; Regulation; Role; S Phase; Saccharomycetales; Signal Pathway; Signal Transduction; Sirolimus; Temperature; Testing; Translational Activation; United States; Virulence; Work; Yeasts; cell type; experimental study; fungus; genetic analysis; genetic approach; mutant; oral infection; oropharyngeal thrush; public health relevance; response; reverse genetics

DESCRIPTION (provided by applicant): The objective of this proposal is to identify the mechanisms which differ between C. albicans yeast and hyphae, for control of proliferation, and for the response to nutritional signaling through the target of rapamycin pathway. In previous work we identified a regulator of hypha-to-yeast morphogenesis, Pes1. Whereas homologs of this gene are essential in all other eukaryotes studied, C. albicans hyphae continue to grow robustly during its depletion, while yeast arrest growth. Our preliminary data show that PES1-depleted hyphae respond to inhibition of Tor nutritional signaling similarly to the wild type, whil yeast fail to arrest growth appropriately. Pes1 is therefore required both for normal yeast proliferation, and for the normal response of yeast to Tor inhibition. We found Pes1 to be required for normal levels of a marker of translational activity which we adapted for use in C. albicans. We also found it to be required for progression of the cell cycle from G1 to S phase. As Pes1 localization differs between cell types and between active and inhibited Tor signaling states, we reason that some of its protein interaction partners differ between these distinct conditions, and preliminary affinity purification experiments support this idea. We aim to identify Pes1 interacting proteins by affinity purification from the distinct cell types and nutritional signaling states. To test the biological relevance of Pes1 interaction partners specific to these conditions, co- immunoprecipitation experiments with reciprocal epitope and affinity tags will be performed, and co- localization of these proteins with Pes1 in the cognate conditions will be confirmed. The role of these Pes1 interaction partners for cell-type specific proliferation control will be investigated by reverse genetic approaches. Localization of Pes1 in yeast and hyphae, and in Tor-active versus -inhibited cells will be determined using co-localization with known markers of specific organelles. Genetic analysis of central nodes of the Tor signaling pathway will answer the question whether cell-type specific signaling is intrinsic to all branches of the pathway or is specific to one branch, and will determine the epistatic and functional relationship of PES1 to these critical elements of Tor nutritional signaling. The results of this work will promote our understanding of specific proliferation control of the ubiquitous C. albicans cell type, the yeast.

描述（由申请人提供）：本提案的目的是确定白色念珠菌酵母和菌丝之间的不同机制，以控制增殖，并通过雷帕霉素途径的靶标响应营养信号。在之前的工作中，我们鉴定了菌丝到酵母形态发生的调节因子 Pes1。尽管该基因的同源物在所研究的所有其他真核生物中都是必需的，但白色念珠菌菌丝在其耗尽期间继续强劲生长，而酵母则抑制生长。我们的初步数据表明，PES1耗尽的菌丝对Tor营养信号的抑制的反应与野生型相似，而酵母无法适当地抑制生长。因此，正常酵母增殖和酵母对 Tor 抑制的正常反应都需要 Pes1。我们发现 Pes1 是翻译活性标记正常水平所必需的，我们将其用于白色念珠菌。我们还发现它是细胞周期从 G1 期进展到 S 期所必需的。由于 Pes1 定位在细胞类型之间以及活跃和抑制的 Tor 信号传导状态之间存在差异，我们推断其一些蛋白质相互作用伙伴在这些不同的条件下有所不同，并且初步的亲和纯化实验支持了这一想法。我们的目标是确定 通过亲和纯化从不同的细胞类型和营养信号状态中提取 Pes1 相互作用蛋白。为了测试特定于这些条件的 Pes1 相互作用配偶体的生物学相关性，将使用相互表位和亲和标签进行共免疫沉淀实验，并确认这些蛋白质与 Pes1 在同源条件下的共定位。这些 Pes1 相互作用伙伴在细胞类型特异性增殖控制中的作用 将通过反向遗传学方法进行研究。 Pes1 在酵母和菌丝中以及在 Tor 活性与抑制细胞中的定位将通过与特定细胞器的已知标记物的共定位来确定。 Tor 信号通路中心节点的遗传分析将回答细胞类型特异性信号传导是该通路所有分支所固有的还是特定于一个分支的问题，并将确定 PES1 与 Tor 营养信号传导这些关键要素的上位和功能关系。这项工作的结果将促进我们对普遍存在的白色念珠菌细胞类型（酵母）的特异性增殖控制的理解。