Regulation of gene expression and genome organization by small RNAs

小RNA调控基因表达和基因组组织

• 批准号: 9330900
• 负责人: Carolyn Marie Phillips
• 金额: $ 41.25万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9330900
• 关键词: Address; Affect; Biological Models; Caenorhabditis elegans; Cell Nucleus; Complex; Congenital Abnormality; Cytology; DNA; Development; Disease; Double-Stranded RNA; Eukaryota; Fire - disasters; Gene Expression; Gene Expression Regulation; Gene Silencing; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Genome Stability; Goals; Human; Human Development; Infertility; Length; Light; Malignant Neoplasms; Messenger RNA; MicroRNAs; Modification; Molecular; Molecular Biology; Movement; Nematoda; Nucleotides; Organism; Pathway interactions; Phylogeny; Play; Positioning Attribute; Process; Protein Family; Proteins; Proteomics; RNA; RNA Interference; Recruitment Activity; Regulator Genes; Research; Role; Route; Set protein; Small RNA; Transcript; Translations; Viral; Work; Yeasts; antiviral immunity; biochemical tools; genome integrity; human disease; mRNA Stability; member; programs

Project Summary: Small RNAs regulate gene expression in a wide range of organisms from yeasts to humans and the fundamental mechanisms governing RNA silencing are conserved across most eukaryotes. The term “small RNA” encompasses a diverse set of small regulatory RNAs, ranging from ~18-30 nucleotides in length, all bound by members of the Argonaute protein family. The first small RNAs identified were microRNAs, discovered in C. elegans as regulators of development timing (Ambros and Ruvkun labs). Several years later, the phenomenon of RNA interference (RNAi), gene silencing induced by the introduction of double stranded RNA, was also discovered in C. elegans (Fire and Mello labs). Much work since then has uncovered the roles of a diverse set of proteins that regulate genes by way of small RNAs at the level of transcription, translation, and mRNA stability. By regulating both endogenous and foreign RNAs, small RNAs play a critical role in development, genome stability, and viral defense. The goal of this research program is to investigate the mechanisms by which RNA silencing pathways modulate gene expression and maintain genome integrity. Like Ambros, Ruvkun, Fire, and Mello, we will use the nematode, C. elegans, as a model system to study RNA silencing because worms combine the best of genetic, cytological, molecular, and biochemical tools. There are many fundamental unanswered questions in our understanding of RNA silencing that we plan to address in this proposal. We and others have determined the localization for many factors in the RNA silencing pathway but there has been little to no cytological exploration the RNA and DNA components. Furthermore, the mechanism by which targeted transcripts are recognized and routed into the RNA silencing pathways is unknown. Our first goal is to localize the mRNAs targeted by RNA silencing and to determine the protein requirements for their physical movement from transcription in the nucleus to turnover in the cytoplasmic RNA silencing compartment, the Mutator foci. We will further examine whether RNA silencing affects the subnuclear positioning of the targeted gene loci, to promote mRNA localization and RNA silencing. Despite much work in the field, there are proteins attributed to small RNA pathway for which the mechanism of action is unknown. We also believe there are components of the pathway that have yet to be identified. Our second goal is to use a combination of molecular biology and cytology to examine uncharacterized components of the pathway in the processes of recruitment and retention of RNA in the Mutator foci and in modification of mRNA targets. Our third goal is to identify and characterize new components of the RNA silencing complex using proteomics. This research will shed light on conserved pathways that regulate gene expression and are critical for development, antiviral immunity, and genome organization.

项目摘要：小 RNA 调节从酵母到多种生物体的基因表达 人类和控制 RNA 沉默的基本机制在大多数真核生物中都是保守的。 术语“小 RNA”涵盖多种小调节 RNA，范围为约 18-30 个核苷酸 长度上，全部与 Argonaute 蛋白家族的成员结合。 microRNA，在线虫中发现作为发育时间的调节剂（Ambros 和 Ruvkun 实验室）。 后来，出现了RNA干扰（RNAi）现象，即由于引入双重基因而引起的基因沉默。 链状 RNA 也在秀丽隐杆线虫中被发现（Fire 和 Mello 实验室）。 在转录水平上通过小RNA调节基因的多种蛋白质的作用， 通过调节内源性和外源性 RNA，小 RNA 发挥着关键作用。 在发育、基因组稳定性和病毒防御中发挥作用。 该研究计划的目标是研究 RNA 沉默途径的机制 像 Ambros、Ruvkun、Fire 和 Mello 一样，我们将使用调节基因表达并保持基因组完整性。 线虫，线虫，作为研究 RNA 沉默的模型系统，因为线虫结合了最好的 遗传、细胞学、分子和生化工具中有许多基本的未解答的问题。 我们计划在本提案中讨论我们对 RNA 沉默的理解。 我们和其他人已经确定了 RNA 沉默途径中许多因素的定位，但是 此外，对RNA和DNA成分的细胞学探索很少甚至没有。 哪些目标转录本被识别并进入 RNA 沉默途径尚不清楚。 目标是定位 RNA 沉默所针对的 mRNA，并确定其所需的蛋白质 从细胞核转录到细胞质 RNA 沉默转换的物理运动 我们将进一步检查 RNA 沉默是否影响亚核。 定位靶基因位点，促进 mRNA 定位和 RNA 沉默。 尽管该领域做了很多工作，但仍有一些蛋白质归因于小 RNA 途径，其中 我们还认为该途径的某些组成部分尚未确定。 我们的第二个目标是结合分子生物学和细胞学进行检查。 RNA募集和保留过程中途径的未表征成分 我们的第三个目标是识别和表征新的突变焦点和 mRNA 靶标。 这项研究将利用蛋白质组学研究 RNA 沉默复合物的组成部分。 调节基因表达并对发育、抗病毒免疫和基因组至关重要的途径 组织。