Regulation of rRNA transcription in mammalian tissues

哺乳动物组织中 rRNA 转录的调控

• 批准号: 10680404
• 负责人: Vikram R. Paralkar
• 金额: $ 40.58万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-21 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10680404
• 关键词: Binding; Biogenesis; Biological Models; Biology; Cells; Chromatin; Complex; DNA Binding; Development; Disease; Energy consumption; Epigenetic Process; Eukaryota; Functional disorder; Genetic Transcription; Goals; Health; Homeostasis; Human; In Vitro; Knowledge; Malignant Neoplasms; Mammals; Maps; Molecular Machines; Mus; Mutation; Normal Cell; Normal tissue morphology; Organ; Organism; Process; Production; Proteins; Proteomics; RNA; Regulation; Reporting; Ribosomal DNA; Ribosomal RNA; Ribosomes; Role; Tissues; Work; cell type; genetic regulatory protein; in vivo; mouse model; novel; ribosomopathy; transcription factor

Abstract / Project Summary Ribosomal RNA (rRNA) comprises 90% of cellular RNA, and ribosome biogenesis is one of the most energy-consuming processes in the cell. The core rRNA transcriptional machinery is evolutionarily ancient and highly conserved from unicellular eukaryotes to mammals, but the bodies of higher organisms have different ribosome production rates in different cell types, responsive to unique tissue-specific demands. Mutations in ribosome biogenesis proteins cause cell-type-specific “ribosomopathies” in humans, manifested by developmental abnormalities, specific organ dysfunctions, or cancers. However, there is little understanding of the transcriptional and epigenetic factors that differentially regulate ribosome biogenesis across normal cells types in intact organisms. Specifically, no one has characterized the protein composition of nucleoli, or the components of rRNA transcription complexes, in any primary mammalian tissue. This represents a key knowledge gap in our understanding of eukaryotic biology. Using quantitative proteomics and transcription factor (TF) mapping studies in a mouse model system, we have identified nucleolar localization and abundant, specific binding to ribosomal DNA (rDNA) of several cell-type-specific TFs (Pu.1, Irf8, Etv6) that are known to be critical for normal development and survival, but whose roles in ribosome biogenesis have never been reported. We propose in this application a combination of unbiased as well as focused approaches to identify and dissect the roles of cell-type-specific rRNA regulators in tissue homeostasis. We will pursue this goal through the following projects: PROJECT 1: DISCOVERY: We will use nucleolar and rDNA-chromatin proteomics in defined primary mouse cell types to identify proteins with cell-type-specific nucleolar localization and rDNA binding. The goal of this project is to identify novel regulators of differential rRNA transcription in intact tissues. PROJECT 2: MECHANISM: We will use ​in vitro and ​in vivo degron and chromatin tethering approaches to dissect the direct roles of rDNA-binding cell-type-specific TFs (Pu.1, Irf8, Etv6, others) in the regulation of rDNA chromatin, rRNA transcription, and tissue homeostasis. The goal of this project is to understand how TF-rDNA binding regulates normal tissue biology. The long-term goal of this work is to gain a detailed understanding of how the ancient process of ribosome biogenesis has evolved to meet diverse tissue needs in complex organisms, and how disruption of this regulation can derange tissue homeostasis and cause disease.

摘要/项目摘要 核糖体 RNA (rRNA) 占细胞 RNA 的 90%，核糖体生物合成是最重要的生物发生之一。 细胞中的核心 rRNA 转录机制在进化上是古老且古老的。 从单细胞真核生物到哺乳动物都高度保守，但高等生物的机体却有不同 不同细胞类型的核糖体生产率，响应独特的组织特异性突变的需求。 核糖体生物发生蛋白在人类中引起细胞类型特异性“核糖体病”，表现为 然而，人们对发育异常、特定器官功能障碍或癌症知之甚少。 差异调节正常细胞核糖体生物发生的转录和表观遗传因子 具体来说，没有人能够表征完整生物体的蛋白质组成或核仁的蛋白质组成。 rRNA 转录复合物的组成部分，在任何初级哺乳动物组织中都是一个关键。 我们对真核生物学理解的知识差距。 通过在小鼠模型系统中进行定量蛋白质组学和转录因子 (TF) 作图研究，我们 已经确定了几种核仁定位以及与核糖体 DNA (rDNA) 的丰富、特异性结合 已知对正常发育和生存至关重要的细胞类型特异性 TF（Pu.1、Irf8、Etv6），但是 其在核糖体生物合成中的作用从未被报道过。 采用公正且有针对性的方法来识别和剖析细胞类型特异性 rRNA 调节因子的作用 我们将通过以下项目来实现这一目标： 项目 1：发现：我们将在确定的原代小鼠中使用核仁和 rDNA 染色质蛋白质组学 细胞类型来识别具有细胞类型特异性核仁定位和 rDNA 结合的蛋白质。 该项目的目的是鉴定完整组织中差异 rRNA 转录的新调节因子。 项目 2：机制：我们将使用体外和体内降解决定子和染色质束缚方法 剖析 rDNA 结合细胞类型特异性 TF（Pu.1、Irf8、Etv6 等）在调节中的直接作用 rDNA 染色质、rRNA 转录和组织稳态 该项目的目标是了解如何实现。 TF-rDNA 结合调节正常组织生物学。 这项工作的长期目标是详细了解核糖体的古老过程如何 生物发生已经进化到满足复杂生物体的不同组织需求，以及如何破坏这种组织 调节会扰乱组织稳态并导致疾病。