The role of phase separation in a transcriptional repression mechanism relevant to Candida albicanspathogenesis.

相分离在与白色念珠菌发病机制相关的转录抑制机制中的作用。

• 批准号: 10607437
• 负责人: Maureen Dowell
• 金额: $ 4.76万
• 依托单位: BROWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10607437
• 关键词: Address; Attenuated; Biochemical; Biochemistry; Biology; C-terminal; Candida; Candida albicans; Cell Nucleolus; Cell Nucleus; Cell model; Cells; Cellular Morphology; Cellular Structures; Cellular biology; Clinical; Complex; Data; Disease; Disseminated candidiasis; Environment; Epigenetic Process; Epithelial; Equilibrium; Eukaryota; Eukaryotic Cell; Exhibits; Fellowship; Gastrointestinal tract structure; Gene Expression; Genetic Transcription; Goals; Heritability; Human; Human Microbiome; Immunologic Deficiency Syndromes; In Vitro; Individual; Infection; Infection prevention; Intervention; Invaded; Lead; Leukocytes; Liquid substance; Location; Mammalian Cell; Microbial Biofilms; Modeling; Molecular Biology; Monitor; Morphology; Mucous Membrane; Mus; Mutation; Mycoses; Oral; Oral mucous membrane structure; Organism; Partner in relationship; Pathogenesis; Phase; Phenotype; Play; Process; Property; Proteins; RNA Polymerase II; RNA Polymerase III; Regulation; Repressor Proteins; Research; Research Personnel; Role; Skin; System; Systemic disease; Systemic infection; Testing; Tissues; Training; Transcription Coactivator; Transcriptional Regulation; Vagina; Virulence; Work; Yeasts; activating transcription factor; career; cell type; experimental study; fly; fungus; gene repression; genetic corepressor; macromolecule; meetings; microbiome; mortality; mouse model; mutant; opportunistic pathogen; oral infection; oropharyngeal thrush; pathogenic fungus; prion-like; programs; recruit; skills; stress granule; symposium; tissue tropism; transcription factor; transcriptome sequencing

Project Summary: Candida albicans is an opportunistic fungal pathogen capable of causing mucosal and systemic disease. Oropharyngeal candidiasis (OPC) is a frequent fungal disease in humans, and can potentially spread to other locations in the body causing systemic infections. Even with clinical intervention, systemic candidiasis has a mortality rate of ~40%. OPC involves the formation of biofilms in the oral mucosa that contain multiple C. albicans morphotypes, including both yeast and hyphal cells. The ability of C. albicans to form different cell types is crucial for virulence and contributes to its capacity to colonize different host tissues. The transition between yeast and hyphal cells is regulated by a network of transcription factors (TFs) that include the co-repressor Ssn6. Deletion of Ssn6 blocks formation of hyphal cells and attenuates C. albicans virulence. Ssn6 is also integral to a second TF network that regulates transitions between ‘white’ and ‘opaque’ states that exhibit different tissue tropisms in the host. Importantly, the Bennett lab has uncovered that multiple TFs (including Ssn6) in both networks contain prion-like domains (PrLDs) and that these can promote phase separation of the corresponding proteins. Phase separation refers to the process of liquid demixing that can generate protein condensates with liquid-like properties. This phenomenon is now implicated in the formation of many cellular structures including nucleoli, Cajal bodies, stress granules, and transcriptional complexes. While a number of studies on TF phase separation have been conducted, these studies primarily focus on transcriptional activators and not on transcriptional co-repressors such as Ssn6. Previous work in our lab has shown that C. albicans TFs containing PrLDs readily form phase-separated condensates when purified or when expressed in mammalian cells. Moreover, deletion or mutation of PrLDs can eliminate the ability of these TFs to function in C. albicans cells. My preliminary data shows Ssn6 is also able to form condensates, both individually and with other fungal TFs. I hypothesize that phase separation plays a key role in the ability of Ssn6 to act as a co-repressor in regulating cell morphology, and that it will contribute to virulence during OPC. In Aim 1, I will examine the biochemical properties of Ssn6 condensates. In Aim 2, I will address how changes to Ssn6 properties impact C. albicans morphotypes in vitro as well as virulence using a mouse model of OPC. Additionally, the goals of this fellowship include developing skill sets in order to lead a successful research career. Attending conferences, presenting to the department and graduate program, and regular meetings with the sponsor are planned to this end. The training plan will be carried out in an environment that benefits from the Molecular Biology, Cell Biology, and Biochemistry Graduate Program. Completion of the proposed goals will advance both the field of Candida biology and the ability of the recipient to become an effective independent scientific researcher.

项目摘要：白色念珠菌是一种机会性真菌病原体，能够引起粘膜和 系统性疾病。口咽部念珠菌病(OPC)是人类常见的真菌病，可能 扩散到身体其他部位，引起全身性感染。即使有临床干预，系统性的 念珠菌病的死亡率约为40%。OPC涉及口腔粘膜中生物膜的形成，其中包含 多种形态的白色念珠菌，包括酵母和菌丝细胞。白色念珠菌形成的能力 不同的细胞类型对毒力至关重要，并有助于其在不同宿主组织中的定植能力。 酵母和菌丝细胞之间的转换受转录因子(TF)网络的调节，该网络 包括共抑制因子Ssn6。SSN6的缺失阻止了菌丝细胞的形成并抑制了白色念珠菌 致命性。SSN6也是第二个Tf网络的组成部分，该网络调节“白色”和“不透明”之间的转换 在宿主中表现出不同组织取向的状态。重要的是，班尼特实验室发现了 两个网络中的TF(包括SSN6)都包含类病毒结构域(PrLD)，这些结构域可以促进阶段 分离出相应的蛋白质。相分离是指液体分离的过程，它可以 产生具有液体性质的蛋白质凝聚体。这一现象现在被认为与 许多细胞结构，包括核仁、卡哈尔小体、应激颗粒和转录复合体。而当 目前已经对转铁蛋白的相分离进行了一些研究，这些研究主要集中在 转录激活子，而不是转录共抑制子，如SSN6。 我们实验室以前的工作表明，含有PrLD的白色念珠菌TF很容易形成相分离 提纯或在哺乳动物细胞中表达时的缩合物。此外，PrLD的缺失或突变可能 消除这些因子在白念珠菌细胞中发挥作用的能力。我的初步数据显示SSN6也能够 形成冷凝物，包括单独的和与其他真菌TF一起形成的。我假设相分离起着关键作用。 在SNN6作为调节细胞形态的辅助抑制因子的能力中的作用，以及它将有助于 OPC期间的毒力。在目标1中，我将研究Sn6冷凝物的生化性质。在《目标2》中，我会 解决SSN6特性的变化如何影响白念珠菌的体外形态类型以及毒力 OPC小鼠模型。 此外，该奖学金的目标包括开发技能集以领导成功的研究 职业生涯。参加会议，向系和研究生课程做报告，并定期与 赞助商计划为此而努力。培训计划将在一个受益于 分子生物学、细胞生物学和生物化学研究生课程。完成拟议的目标将 促进假丝酵母菌生物学领域和接受者成为有效独立的能力 科学研究人员。