Defining and Controlling Protein-RNA interactions in editing and interference pathways

定义和控制编辑和干扰途径中蛋白质-RNA 相互作用

• 批准号: 10390419
• 负责人: PETER A. BEAL
• 金额: $ 54.78万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10390419
• 关键词: ADAR1; Address; Adenosine; Antisense Oligonucleotides; Area; Autoimmune Diseases; Award; Bees; Chemicals; Chemistry; Codon Nucleotides; Complex; Coupled; DRADA2b protein; Development; Disease; Enzymes; Gene Silencing Pathway; Genes; Guanosine; Human; Inosine; Laboratories; Lead; Length; Malignant Neoplasms; Methods; MicroRNAs; Modification; Molecular Biology; Mutation; Pathway interactions; Process; Property; Proteins; RNA; RNA Editing; RNA Interference Pathway; RNA Sequences; RNA-Binding Proteins; RNA-Protein Interaction; Reagent; Regulation; Research; Research Personnel; Research Support; Small Interfering RNA; Structure; Therapeutic; Up-Regulation; Work; design; gene function; human disease; improved; inhibitor; interest; mutant; novel; novel therapeutics; pseudotoxoplasmosis syndrome; screening; skin disorder; targeted cancer therapy; therapeutic RNA; tool

This Maximizing Investigators Research Award (MIRA) application is proposed to support research in the Beal lab at UC Davis focused on defining and controlling protein-RNA interactions in RNA editing and RNA interference pathways. The RNA editing ADAR enzymes convert adenosines (A) to inosines (I) in duplex RNA. Since I can behave similarly to guanosine (G) in RNA, this modification can have profound effects on the structure and function of the modified RNA including, but not limited to, changes in the meaning of specific codons (recoding). Mutations in the human ADAR1 gene cause the skin disorder Dyschromatosis Symmetrica Hereditaria (DSH) and the autoimmune disease Aicardi- Goutieres Syndrome (AGS). Also, ADAR1 upregulation and hyper editing has been observed in several different cancers. Despite the significance of this form of regulation of RNA structure and function, there remain key gaps in our understanding of A to I RNA editing. In addition, given ADARs’ ability to change RNA sequence, there is growing interest in harnessing this property and directing it to correct disease-associated G-to-A mutations. Key questions in this field that will be addressed in this project are: 1) What are the structures of key protein-RNA complexes in editing pathways? Structures of full length human ADAR2 bound to different RNA substrates along with structures of ADAR1 bound to RNA are necessary for a full understanding of substrate recognition and selectivity in RNA editing. 2) Can we develop potent, selective and low MW ADAR inhibitors? Such inhibitors could serve as lead compounds in the development of ADAR1-targeted cancer therapies. 3) Can we develop new strategies to evolve mutant editing enzymes and novel substrate RNAs? The results of these efforts will inform the design of highly efficient and selective reagents for directed RNA editing applications. Our laboratory also has a long standing interest in the development of chemical modifications of RNA that can control the interaction with RNA-binding proteins. Much of our recent work in this area has focused on controlling the interaction of RNA with components of siRNA-triggered or miRNA- triggered gene silencing pathways. The use of the RNAi pathway to study gene function has become a powerful tool in molecular biology and has been exploited in the development of new therapeutics. However, specific issues exist that limit its application. These issues include off-target effects that arise from the ability of an siRNA guide strand to function as a miRNA. In addition, antisense oligonucleotides targeting miRNAs (anti-miRs) have significant therapeutic potential and require chemical modification for stability and efficacy. Up to this point, the development of new chemical modifications of therapeutic RNAs has been largely an ad hoc process. The key question addressed in this aspect of the proposed project is: Can we develop an effective systematic approach to new RNA modifications that modulate protein-RNA interactions in interference pathways? The immediate impact of these studies will be to provide new modifications to siRNAs and anti-miRs that improve potency and selectivity. However, our continued refinement of an approach that uses computational screening coupled with versatile RNA modification chemistry will be generally applicable other projects that involve chemically modified RNA for therapeutics.

这个最大化研究者研究奖（MIRA）的应用程序，建议支持研究， 加州大学戴维斯分校的比尔实验室专注于定义和控制RNA编辑中的蛋白质-RNA相互作用 和RNA干扰途径。RNA编辑阿达尔酶将腺苷（A）转化为肌苷 (I)在双链RNA中。由于I的行为类似于RNA中的鸟苷（G），因此这种修饰可以具有 对修饰的RNA的结构和功能的深远影响，包括，但不限于， 改变特定密码子的含义（重新编码）。人类ADAR 1基因突变导致 遗传性对称性色素异常症（DSH）和自身免疫性疾病Abladii- Goutieres综合征（AGS）。此外，在小鼠中观察到ADAR 1上调和超编辑。 几种不同的癌症尽管这种形式的RNA结构调节的重要性， 尽管如此，我们对A到I RNA编辑的理解仍然存在关键差距。此外，鉴于ADAR的 改变RNA序列的能力，人们越来越有兴趣利用这种特性并指导它 来纠正疾病相关的G转A突变在这一领域的关键问题，将在 1）编辑途径中的关键蛋白质-RNA复合物的结构是什么？ 与不同RNA底物结合的全长人ADAR 2的结构与以下结构沿着： 与RNA结合的ADAR 1对于全面了解底物识别和选择性是必要的 在RNA编辑中。2)我们能否开发出有效的、选择性的和低分子量的阿达尔抑制剂？此类抑制剂 可以作为开发ADAR 1靶向癌症疗法的先导化合物。3)我们能 开发新的策略来进化突变编辑酶和新的底物RNA？的 这些努力的结果将为设计高效和选择性的定向RNA试剂提供信息 编辑应用程序。 我们的实验室也有一个长期的兴趣，在化学修饰的发展， 可以控制与RNA结合蛋白相互作用的RNA。我们最近在这方面的工作 一直专注于控制RNA与siRNA触发的或miRNA触发的组分的相互作用， 引发了基因沉默途径利用RNAi途径研究基因功能已成为 它是分子生物学中的一种强大工具，并已被用于开发新的治疗方法。 然而，存在一些具体问题，限制了其适用。这些问题包括脱靶效应， 这源于siRNA引导链作为miRNA发挥功能的能力。此外，反义 靶向miRNAs的寡核苷酸（抗miRs）具有显著的治疗潜力， 化学修饰以提高稳定性和功效。到目前为止，新化学品的开发 治疗性RNA的修饰在很大程度上是一个特别的过程。解决的关键问题 在这方面的拟议项目是：我们能否制定一个有效的系统方法，以新的 在干扰途径中调节蛋白质-RNA相互作用的RNA修饰？的 这些研究的直接影响将是提供对siRNA和抗miR的新修饰， 提高效力和选择性。然而，我们不断改进的方法， 计算机筛选与通用RNA修饰化学相结合， 适用于其他涉及用于治疗的化学修饰RNA的项目。