DNA Repair Mechanisms in Candida albicans

白色念珠菌的 DNA 修复机制

• 批准号: 6695813
• 负责人: Richard Arthur Calderone
• 金额: $ 16.85万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2006-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6695813
• 关键词: Candida albicans; DNA repair; SDS polyacrylamide gel electrophoresis; candidiasis; fungal genetics; fungal proteins; gene expression; genetic recombination; histogenesis; laboratory mouse; laboratory rat; microarray technology; mutant; polymerase chain reaction; protein structure function; virulence; western blottings

DESCRIPTION (provided by applicant): DNA repair has not been studied in the human pathogen Candida albicans in spite of numerous observations that suggest extensive genomic variability among clinical isolates. This variability has been associated with a variety of genomic alterations that include, deletions, translocations, ploidy changes, as well as loss of chromosomes or parts of chromosomes. The relationship of these changes to adaptation, virulence, and growth of the fungus in the host is uncertain, but sugar utilization and drug resistance have been correlated in vitro with aneuloploidy. Homologous recombination [HR] and non-homologous end-joining [NHEJ] should be important components in DNA repair to ensure the fidelity of these genomic alterations. Our preliminary studies with Rad52p of the HR and Lig4p of the NHEJ pathways suggest that both proteins provide growth functions in addition to DNA repair in C. albicans. This fact along with the surprising genomic variability of this pathogen suggest differences between the DNA repair pathways of S. cerevisiae and C. albicans. To understand the roles of HR and NHEJ in this organism, we propose four specific aims. Specific aim 1 includes the construction of new mutants in each of the two pathways, HDF1 in NHEJ and RAD59 in HR. In specific aim 2, we will complete our analysis of PIF1 and RRM3, two proteins identified from a two-hybrid screen that interact with Lig4p. The role of HR and NHEJ pathway proteins in DNA repair and morphogenesis is the focus of specific aim 3. In addition, microarray analysis will be used to identify up and down regulated genes in selected mutants of C. albicans when compared to wild type cells. Finally, aim 4 is designed to evaluate the role of several gene deleted mutants in the pathogenesis of candidiasis. We propose to use three mouse models of candidiasis to evaluate virulence, including, invasive, oral, and vaginal models so as to identify a site-specific role of these genes in virulence. In summary, our studies will provide critical information on the contribution of DNA repair to the adaptation, growth, variability and virulence of C. albicans.

描述（由申请人提供）： 尽管大量观察表明临床分离株之间存在广泛的基因组变异，但尚未在人类病原体白色念珠菌中研究 DNA 修复。这种变异性与多种基因组改变有关，包括缺失、易位、倍性变化以及染色体或部分染色体的丢失。这些变化与宿主真菌的适应、毒力和生长的关系尚不确定，但糖利用和耐药性在体外与非整倍体相关。同源重组 [HR] 和非同源末端连接 [NHEJ] 应该是 DNA 修复的重要组成部分，以确保这些基因组改变的保真度。我们对 HR 通路的 Rad52p 和 NHEJ 通路的 Lig4p 的初步研究表明，这两种蛋白在白色念珠菌中除了 DNA 修复之外还提供生长功能。这一事实以及该病原体令人惊讶的基因组变异表明酿酒酵母和白色念珠菌的 DNA 修复途径之间存在差异。为了了解 HR 和 NHEJ 在该有机体中的作用，我们提出了四个具体目标。具体目标 1 包括在两条途径中构建新突变体，即 NHEJ 中的 HDF1 和 HR 中的 RAD59。在具体目标 2 中，我们将完成对 PIF1 和 RRM3 的分析，这两种蛋白质是通过双杂交筛选鉴定出的，与 Lig4p 相互作用。 HR 和 NHEJ 途径蛋白在 DNA 修复和形态发生中的作用是具体目标 3 的重点。此外，微阵列分析将用于识别与野生型细胞相比，白色念珠菌选定突变体中上调和下调的基因。最后，目标 4 旨在评估几种基因缺失突变体在念珠菌病发病机制中的作用。我们建议使用三种念珠菌病小鼠模型来评估毒力，包括侵入性、口腔和阴道模型，以便确定这些基因在毒力中的位点特异性作用。总之，我们的研究将提供关于 DNA 修复对白色念珠菌的适应、生长、变异和毒力的贡献的关键信息。