The identification of fundamental molecular regulators of ribosomal DNA and nucleolar organization in fission yeast

裂殖酵母核糖体 DNA 和核仁组织基本分子调节因子的鉴定

• 批准号: 10845906
• 负责人: Alexandria Jane Cockrell
• 金额: $ 3.87万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-03 至 2025-01-02
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10845906
• 关键词: Address; Architecture; Area; Automobile Driving; Bacterial DNA; Biogenesis; Biological Models; Cell Cycle; Cell Cycle Progression; Cell Cycle Regulation; Cell Nucleolus; Cell physiology; Cells; Cellular biology; Chromosome Structures; Chromosomes; Classification; DNA; DNA analysis; DNA-Binding Proteins; Exhibits; Fission Yeast; Fluorescence Microscopy; Future; Genetic; Genetic Transcription; Genome; Goals; Human; Image; Image Analysis; Imaging Device; Interphase; Knowledge; Label; Maintenance; Malignant Neoplasms; Measures; Methods; Modeling; Molecular; Molecular Biology; Molecular Biology Techniques; Morphology; Neurodegenerative Disorders; Nuclear; Nucleolar Organizer Region; Nucleolar Proteins; Organelles; Organism; Physiologic pulse; Process; Production; Protein Biosynthesis; Proteins; Quantitative Microscopy; RNA; RNA Processing; Research; Ribosomal DNA; Ribosomal Proteins; Ribosomal RNA; Ribosomes; Role; Saccharomycetales; Structure; Testing; Transcript; Variant; cell growth; deletion library; field study; genetic approach; genetic manipulation; genome-wide; high throughput analysis; human disease; imaging genetics; innovation; mutant; novel; novel strategies; nucleolar organizing region; protein complex; tool

Project Abstract Ribosome production relies on a nuclear organelle called the nucleolus. Within this structure, ribosomal DNA (rDNA) is transcribed to form RNA transcripts that associate with ribosomal proteins. Nucleolar architecture is altered in many human diseases including numerous cancers, prompting several studies to search for regulators of nucleolar morphology. While these studies rely on analysis of nucleolar proteins or ribosome production to identify molecular regulators, few studies have defined cell cycle-specific mechanisms for regulating nucleolar structure. Furthermore, no studies have examined the impact of rDNA spatial organization on nucleolar morphology despite rDNA loci’s known role as Nucleolar Organizer Regions. The long-term goal is to understand conserved regulatory mechanisms of rDNA and nucleolar organization. The overall objectives of this proposal are to (i) to identify molecular regulators of rDNA spatial organization and (ii) define the processes driving cell cycle-regulated nucleolar morphology. The central hypothesis is that the spatial organization of rDNA is regulated, in part, by chromosome organizing proteins and ribosome biogenesis; furthermore, these ribosome biogenesis processes are cell cycle-regulated, driving dynamic reorganization of nucleolar morphology during interphase. The rationale for this study is that identification of conserved molecular regulators of rDNA and nucleolar organization in fission yeast will provide a template for future research in higher organisms. This central hypothesis will be tested by two specific aims: 1) identify molecular regulators of rDNA spatial organization in fission yeast; and 2) define the processes driving cell cycle-regulated nucleolar morphology. For aim 1, a novel tool for analysis of rDNA spatial organization in live cells has been developed in fission yeast. This tool will be used to quantify rDNA spatial organization by fluorescence microscopy in candidate mutants with altered chromosome organization and DNA topology factors. This analysis will be expanded by a genome-wide high-throughput imaging screen to broadly identify regulators of rDNA spatial organization. Aim 2 will apply fluorescence microscopy, cell biology, and molecular biology approaches to examine the role of cell cycle-regulated ribosome biogenesis factors in interphase nucleolar morphology. These studies examine rDNA and nucleolar morphology in fission yeast, a model system notable for its application to higher organisms and ease of genetic manipulation. To understand the relationship between nucleolar morphology and human disease, the regulatory mechanisms behind rDNA and nucleolar organization must be identified. This study applies innovative imaging tools with advanced cellular and molecular biology techniques to broadly identify fundamental molecular regulators of rDNA and nucleolar morphology, providing a framework for future studies in human cells.

项目摘要 核糖体的产生依赖于称为核olus的核细胞器。在这种结构中，核糖体DNA （rDNA）被转录为形成与核糖体蛋白相关的RNA转录本。核仁结构是 许多人类疾病都改变了包括众多癌症，促使一些研究搜索 核形态的调节剂。尽管这些研究依赖于核蛋白或核糖体的分析 生产以鉴定分子调节剂，很少有研究确定了细胞周期特异性机制 调节核结构。此外，没有研究检查rDNA空间组织的影响 关于核形态任务rDNA基因座的核仁组织者区域的作用。长期目标 是要了解rDNA和核仁组织的配置调节机制。总体目标 该建议的是（i）确定rDNA空间组织的分子调节剂，并（ii）定义 驱动细胞周期调节的核形态的过程。中心假设是空间 rDNA的组织部分通过染色体组织蛋白质和核糖体生物发生来调节。 此外，这些核糖体生物发生过程是细胞周期调节的，推动了动态重组 相间期间的核酸形态。这项研究的理由是鉴定保守的 裂变酵母中rDNA和核仁组织的分子调节剂将为将来提供模板 在较高生物体中进行研究。该中心假设将通过两个特定目的测试：1）确定分子 裂变酵母中rDNA空间组织的调节剂； 2）定义驱动细胞周期调节的过程 核仁形态。对于AIM 1，一种用于分析活细胞中rDNA空间组织的新工具已经是 在裂变酵母中开发。该工具将通过荧光来量化rDNA空间组织 具有改变染色体组织和DNA拓扑因素的候选突变体的显微镜。这 分析将通过全基因组高通量成像屏幕扩展，以广泛识别 rDNA空间组织。 AIM 2将应用荧光显微镜，细胞生物学和分子生物学 检验细胞周期调节的核糖体生物发生因子在相间核仁中的作用的方法 形态学。这些研究检查了裂变酵母中的rDNA和核仁形态，该模型系统著名 它应用于更高的生物体和易于遗传操作。了解关系 在核形态和人类疾病之间，rDNA和核的调节机制 必须确定组织。这项研究应用了具有高级蜂窝和的创新成像工具 分子生物学技术可广泛识别rDNA和核仁的基本分子调节剂 形态学，为人类细胞的未来研究提供了一个框架。