Center for dynamic RNA epitranscriptomes - Renewal

动态 RNA 表观转录组中心 - 更新

• 批准号: 10159493
• 负责人: CHUAN HE
• 金额: $ 292.45万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-27 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10159493
• 关键词: Adenosine; Affect; Animal Model; Area; Basic Science; Binding Sites; Biological; Biological Process; Biology; Cell Differentiation process; Chemicals; Chemistry; Chromatin; Chromosomes; Clinical; Communities; Complex; Databases; Development; Disease; Engineering; Enhancers; Enzymes; Evolution; Gene Expression Regulation; Genetic Transcription; Genomics; Introns; Investigation; Life; Mammals; Maps; Medical; Messenger RNA; Methods; Methylation; Modification; Mutation; Neurons; Nucleic Acids; Physiological; Play; Post-Transcriptional RNA Processing; Property; Proteins; Protocols documentation; Pseudouridine; RNA; RNA Processing; RNA metabolism; RNA-Binding Proteins; Reader; Regulation; Research; Resolution; Reverse Transcription; Ribosomal RNA; Role; Sampling; Site; Study models; System; Testing; Tissues; Transcript; Transfer RNA; Translations; Untranslated RNA; Up-Regulation; Work; base; bioinformatics tool; chemical group; computational pipelines; demethylation; epitranscriptome; epitranscriptomics; experimental study; human disease; method development; nanopore; new technology; promoter; sequencing platform; stem cell differentiation; transcriptome

Project Summary/Abstract Chemical modifications on mammalian messenger RNA (mRNA) have recently been shown to play critical and diverse regulatory roles in mRNA metabolism and translation. For example, the most abundant mRNA modification, N6-methyladenosine (m6A), is crucial for mammalian stem cell differentiation and tissue development in almost all systems tested so far. Dedicated proteins, many of which are essential in mammals, have evolved to install, recognize, and remove m6A marks (writers, readers, and erasers, respectively). Dysregulation of m6A methylation has been connected to a variety of human diseases and disorders. Functional roles have also been proposed for other internal modifications present in mammalian mRNA, including, but not limited to: pseudouridine (Ψ), 5-methylcytosine (m5C), 2’-O-methylation (Nm), N1-methyladenosine (m1A), N7- methylguanosine (m7G), and N3-methylcytosine (m3C). Our most recent research has uncovered modifications on chromosome-associated regulatory RNAs (carRNAs), such as promoter-associated RNA (paRNA), enhancer RNA (eRNA), and repeat RNAs, as well as frequent modifications in introns of pre-mRNA. The carRNA modifications have been shown to regulate chromatin state and transcription, and intron modifications may affect pre-mRNA processing. Despite rapid advances in the discovery and functional characterization of various RNA modifications and their effector proteins, a significant bottleneck limits the entire field of epitranscriptomics research: a dearth of quantitative sequencing methods that can comprehensively map most RNA modifications at base resolution with exact modification fraction information. The availability of such methods is critical for assessing the importance of these modifications in different regions of mRNA, examining the effects of dynamic changes in modification fraction, assigning modifications to different writers and analyzing their functional relevance, identifying target transcripts and sites of demethylation and analyzing their functional relevance, discovering new effectors for RNA modifications by overlapping with known RBP-binding sites or genomics features, and evaluating the physiological consequences of RNA modifications in biological processes. We have established both nucleic acid chemistry and directed protein evolution platforms to invent new technologies that transform RNA modifications to be read out as mutations or deletions that are universally compatible with extant sequencing platforms. Computational pipelines and RNA modification databases will be built to support the new method development and epitranscriptome research in the broad community. These new technologies will be optimized to work on low-input samples, particularly neuronal and clinical samples. We will focus on integrating new methods into robust protocols to map multiple RNA modifications in single experiments. Our proposed research will deliver high-throughput, high-resolution, and high-sensitivity methods that simultaneously map multiple RNA modifications in all biological areas.

项目概要/摘要 最近研究表明，哺乳动物信使 RNA (mRNA) 的化学修饰发挥着至关重要的作用 mRNA 代谢和翻译中的多种调节作用例如，最丰富的 mRNA。 N6-甲基腺苷 (m6A) 修饰对于哺乳动物干细胞分化和组织至关重要 迄今为止，几乎所有测试系统都在开发专用蛋白质，其中许多对哺乳动物来说是必需的， 已经发展到可以安装、识别和删除 m6A 标记（分别为写入器、读取器和擦除器）。 m6A 甲基化失调与多种人类疾病和功能失调有关。 还提出了哺乳动物 mRNA 中存在的其他内部修饰的作用，包括但不限于 仅限于：假尿苷 (Ψ)、5-甲基胞嘧啶 (m5C)、2’-O-甲基化 (Nm)、N1-甲基腺苷 (m1A)、N7- 我们最近的研究发现了甲基鸟苷 (m7G) 和 N3-甲基胞嘧啶 (m3C) 的修饰。 染色体相关调节 RNA (carRNA)，例如启动子相关 RNA (paRNA)、增强子 RNA (eRNA) 和重复 RNA，以及前 mRNA 内含子的频繁修饰。 修饰已被证明可以调节染色质状态和转录，内含子修饰可能会影响 尽管各种 RNA 的发现和功能表征取得了快速进展。 修饰及其效应蛋白，是限制表观转录组学整个领域的一个重要瓶颈 研究：缺乏能够全面绘制大多数 RNA 修饰图谱的定量测序方法 具有精确修饰分数信息的碱基分辨率，此类方法的可用性对于 评估这些修饰在 mRNA 不同区域的重要性，检查动态的影响 修改比例的变化，将修改分配给不同的作者并分析其功能 相关性，识别目标转录本和去甲基化位点并分析其功能相关性， 通过与已知的 RBP 结合位点或基因组重叠来发现 RNA 修饰的新效应 特征，并评估生物过程中 RNA 修饰的生理后果。 建立了核酸化学和定向蛋白质进化平台，以发明新技术 将 RNA 修饰转化为突变或缺失，与现有的普遍兼容 将建立计算管道和RNA修饰数据库来支持新的技术。 这些新技术将在广泛的社区中进行方法开发和表观转录组研究。 针对低输入样本进行优化，特别是神经和临床样本，我们将专注于整合。 我们提出的新方法可用于在单个实验中绘制多个 RNA 修饰的稳健协议。 研究将提供高通量、高分辨率和高灵敏度的方法，同时绘制 所有生物领域的多种 RNA 修饰。