Cdc14 phosphatase - novel roles in drug resistance, virulence, and the response to cell wall stress in fungal pathogens

Cdc14磷酸酶——在真菌病原体的耐药性、毒力和细胞壁应激反应中的新作用

• 批准号: 10657007
• 负责人: MARK C HALL
• 金额: $ 62.27万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-17 至 2027-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10657007
• 关键词: Active Sites; Alleles; Animals; Antifungal Agents; Auxins; Benign; Binding; Biochemical; Biological; Biological Assay; Biological Process; C-terminal; Candida albicans; Candida auris; Candidiasis; Cell Wall; Cell division; Cells; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytokinesis; Data; Defect; Development; Disease; Disease Outbreaks; Drug Targeting; Drug resistance; Enzymes; Evolution; Family; Follow-Up Studies; Fungi Model; Future; Genetic Transcription; Growth; Growth and Development function; Hospitals; Hypersensitivity; Immune; Impairment; Individual; Infection; Life; Link; Maintenance; Methods; Mitotic; Modeling; Molecular; Molecular Target; Multi-Drug Resistance; Mus; Mycoses; Orthologous Gene; Pathogenesis; Pathway interactions; Persons; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Process; Property; Protein phosphatase; Proteomics; Regulation; Resolution; Role; Saccharomyces cerevisiae; Severities; Signal Induction; Signal Transduction; Specificity; Stress; Structure; Substrate Specificity; System; Systemic infection; Tail; Testing; Therapeutic; Virulence; Virulence Factors; Work; World Health; biological adaptation to stress; combat; design; drug development; drug resistant pathogen; drug sensitivity; echinocandin resistance; experimental study; fungus; human pathogen; inhibitor; insight; loss of function; mRNA Translation; mouse model; novel; pathogen; pathogenic fungus; phosphoproteomics; resistance factors; response; transcriptomics; Active Sites; Alleles; Animals; Antifungal Agents; Auxins; Benign; Binding; Biochemical; Biological; Biological Assay; Biological Process; C-terminal; Candida albicans; Candida auris; Candidiasis; Cell Wall; Cell division; Cells; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytokinesis; Data; Defect; Development; Disease; Disease Outbreaks; Drug Targeting; Drug resistance; Enzymes; Evolution; Family; Follow-Up Studies; Fungi Model; Future; Genetic Transcription; Growth; Growth and Development function; Hospitals; Hypersensitivity; Immune; Impairment; Individual; Infection; Life; Link; Maintenance; Methods; Mitotic; Modeling; Molecular; Molecular Target; Multi-Drug Resistance; Mus; Mycoses; Orthologous Gene; Pathogenesis; Pathway interactions; Persons; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Process; Property; Protein phosphatase; Proteomics; Regulation; Resolution; Role; Saccharomyces cerevisiae; Severities; Signal Induction; Signal Transduction; Specificity; Stress; Structure; Substrate Specificity; System; Systemic infection; Tail; Testing; Therapeutic; Virulence; Virulence Factors; Work; World Health; biological adaptation to stress; combat; design; drug development; drug resistant pathogen; drug sensitivity; echinocandin resistance; experimental study; fungus; human pathogen; inhibitor; insight; loss of function; mRNA Translation; mouse model; novel; pathogen; pathogenic fungus; phosphoproteomics; resistance factors; response; transcriptomics

PROJECT SUMMARY Opportunistic fungal infection of immune-compromised individuals is an escalating world health problem. Recent lethal outbreaks of multi drug-resistant Candida auris in hospitals and the rise of drug resistance in normally benign commensal fungi like C. glabrata highlight the severity of the problem. Current treatment options for fungal infections are limited to a few antifungal drug classes that are becoming increasingly ineffective. There is a pressing need for new molecular targets for antifungal development to deal with drug- resistant pathogens. This project will characterize a newly identified C. albicans virulence and drug resistance factor, the Cdc14 protein phosphatase. Our recent work has uncovered novel roles for C. albicans Cdc14 in regulating cell wall integrity, septation, echinocandin sensitivity, and hyphal development, all processes tied to virulence. Importantly, even modest reduction in Cdc14 activity level severely compromises virulence in a mouse model of invasive candidiasis. In contrast, Cdc14 is dispensable for normal development, growth, and cell division in animals. Cdc14 is highly conserved in fungi and its unique and strict active site specificity implies that development of potent and highly selective inhibitors should be achievable, something that has been challenging with other protein phosphatases. Our overall objective is to characterize the mechanisms by which Cdc14 regulates virulence-associated biological processes in C. albicans. In Aim 1 we will characterize Cdc14 regulation of cell wall integrity and septation. In Aim 2 we will characterize Cdc14 regulation of hyphal initiation and maintenance. In Aim 3 we will characterize the mechanisms by which Cdc14 itself is regulated by cell wall stress and hypha-inducing signals. In Aims 1 and 2 we will employ unbiased omics approaches to identify the relevant substrates of Cdc14 and the transcriptional circuits under Cdc14 control. In Aim 1 we will directly characterize the cell wall defects arising from Cdc14-deficiency. We will also test specific models for Cdc14 function in promoting cell wall integrity and hyphal initiation in Aims 1 and 2, respectively. In Aim 3 we will focus on phosphoregulation of the disordered Cdc14 C-terminal tail, which is a hub for integration of regulatory signals in model fungi. We will use quantitative phosphoproteomics to understand the dynamic phosphorylation of C. albicans Cdc14 during cell wall stress, cytokinesis/septation, and initiation of hyphal differentiation. All three aims will conclude with structure-function analyses using biochemical, cell biological, and waxworm and mouse infection assays to characterize the physiological significance of Cdc14 function and phosphoregulation, including the importance for pathogenesis. Collectively, the results will define the molecular mechanisms by which Cdc14 promotes several virulence-related biological processes that will be useful in assessing its future potential as an antifungal target. The identification of Cdc14 substrates and effectors may provide additional candidate antifungal targets. The high conservation of Cdc14 structure, activity, and specificity across the fungal kingdom implies the results will be relevant to many other fungal pathogens.

项目摘要 免疫力低下的个体的机会性真菌感染是一个不断升级的世界卫生问题。 最近在医院中抗药性念珠菌的致命爆发以及耐药性的兴起 通常，良性的共生真菌（如C. glabrata）突出了问题的严重性。当前治疗 真菌感染的选择仅限于一些越来越多的抗真菌药物类别 无效。迫切需要对抗真菌发育的新分子靶标，以应对药物 抗性病原体。该项目将表征新近确定的白色念珠菌毒力和耐药性 因子，Cdc14蛋白磷酸酶。我们最近的工作发现了白色念珠菌CDC14的新作用 调节细胞壁完整性，分隔，echinocandin敏感性和菌丝发育，所有过程与 毒力。重要的是，即使是CDC14活性水平的适度降低也会严重损害A中的毒力 浸润性念珠菌病的小鼠模型。相反，CDC14对于正常发育，增长和 动物的细胞分裂。 CDC14在真菌及其独特而严格的活性场地特异性中高度保守 意味着应该达到有效和高度选择性抑制剂的发展，这是 在其他蛋白质磷酸酶方面充满挑战。我们的总体目标是通过 CDC14调节白色念珠菌中与毒力相关的生物学过程。在AIM 1中，我们将描述 CDC14细胞壁完整性和分离的调节。在AIM 2中，我们将表征CDC14菌丝调节 启动和维护。在AIM 3中，我们将表征CDC14本身调节的机制 细胞壁应力和菌丝诱导信号。在目标1和2中，我们将采用公正的OMIC方法 确定CDC14的相关底物和CDC14对照下的转录电路。在目标1中，我们将 直接表征由CDC14缺陷引起的细胞壁缺陷。我们还将测试特定模型的 Cdc14分别在AIM 1和2中促进细胞壁完整性和菌丝起始中的功能。在目标3中我们 将着重于无序CDC14 C末端尾的磷酸化，这是整合的枢纽 模型真菌中的监管信号。我们将使用定量的磷蛋白组学来了解动态 在细胞壁应力，细胞因子/分离和菌丝的开始期间白色念珠菌CDC14的磷酸化 分化。这三个目标将以生化，细胞生物学，结构功能分析结束 以及蜡虫和小鼠感染分析，以表征Cdc14功能的生理意义和 磷化器调节，包括对发病机理的重要性。总体而言，结果将定义分子 CDC14促进了几种与毒力相关的生物学过程的机制 评估其作为抗真菌目标的未来潜力。 CDC14底物和效应子的识别可能 提供其他候选抗真菌靶标。 CDC14结构，活动和 真菌王国的特异性意味着结果将与许多其他真菌病原体有关。