Mechanisms of Microprocessor Function and Regulation

微处理器功能和调节机制

• 批准号: 10364326
• 负责人: Yunsun Nam
• 金额: $ 36.97万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10364326
• 关键词: Address; Affect; Affinity; Apical; Binding; Biochemical; Biochemistry; Biogenesis; Biological Assay; Biology; Cell Maintenance; Complex; Cryoelectron Microscopy; Defect; Detection; Development; Diabetes Mellitus; Disease; Distal; Double-Stranded RNA; Enzymes; Family; Future; Gene Cluster; Gene Expression; Gene Expression Regulation; Genes; Genome; Goals; Homo; Human; In Vitro; Individual; Knowledge; Left; Libraries; Link; Malignant Neoplasms; Metabolic syndrome; MicroRNAs; Microprocessor; Modeling; Molecular; Play; Process; Proteins; RNA; RNA Sequences; Regulation; Regulator Genes; Research; Rest; Ribonuclease III; Role; SAFB gene; Signal Transduction; Site; Specificity; Stress; Structure; Substrate Specificity; Tertiary Protein Structure; Transcript; Untranslated RNA; Work; X-Ray Crystallography; arm; cellular development; developmental disease; dimer; grasp; helicase; insight; mutant; novel; organ growth; preference; protein expression; recruit; response; stem; stem cells; stoichiometry; success; three dimensional structure

Project Summary/Abstract (30 lines) MicroRNAs (miRNAs) constitute a large family of short, non-coding, regulatory RNAs that modulate protein expression. Abnormal miRNA levels are associated with many diseases, including developmental defects and various cancers. To generate functional miRNAs, primary transcripts (pri-miRNAs) generally need to be first cleaved by an RNaseIII, Drosha. This critical step of miRNA biogenesis needs to be controlled, in both accuracy and efficiency, to maintain proper gene regulation. The processing enzyme Drosha requires its partner protein, DGCR8, for protein stability and substrate specificity. A Drosha molecule binds homo-dimeric DGCR8 to carry out pri-miRNA processing, but higher-order complexes may also form because clustering of pri-miRNAs in the genome enhances processing. Our recent groundbreaking cryo-electron microscopy (cryo- EM) structures provide atomic models of the Microprocessor-pri-miRNA complex in action. Elucidating how the proteins are organized around the RNA stem-loop also revealed how each distal end (basal or apical) is independently recognized but also linked to each other via a molecular ruler connecting the detection modules. The proposed research builds on our previous successes, as the structural framework will enable us to gain novel fundamental insights into how the macromolecular recognition is accomplished. Our overall goal is to understand the recognition of pri-miRNAs by Microprocessor at the molecular level. We hypothesize that RNA structural features and context-dependent sequence preferences dictate the processing fate of each individual pri-miRNA. We will dissect how the recognition is accomplished at each of the basal and apical junctions of pri- miRNAs. We will also investigate how diverse RNA sequences and structures from different pri-miRNAs affect proper recognition at each junction, to reveal the plasticity of the multipart machinery. Our previous work has also left us poised to address urgent questions on how clustering of pri-miRNAs enhances processing by Microprocessor, which is crucial for our overall understanding of miRNA biogenesis and has profound implications for the interpretation of previous and future results in a wide variety of fields obtained by manipulation of miRNA genes. A better grasp of the core recognition mechanisms will help us explain unique targets such as clustered pri-miRNAs. Together, the proposed studies will provide a comprehensive understanding of processing and regulation of miRNAs, important regulators of gene expression. Our work on deciphering how structure affects RNA recognition is a fundamentally important question and likely be insightful for many processes involving structured RNAs beyond miRNAs.

项目摘要/摘要(30行) MicroRNAs(MiRNAs)构成一个短的、非编码的、调节蛋白质的RNAs大家族 表情。MiRNA水平异常与许多疾病有关，包括发育缺陷和 各种癌症。为了产生具有功能的miRNAs，初级转录本(pri-miRNAs)通常需要首先 被RNaseIII，DROSHA切割。MiRNA生物发生的这一关键步骤需要在两者中进行控制 准确性和效率，以保持适当的基因调控。加工酶DROSHA需要其 伙伴蛋白Dgcr8，用于蛋白质稳定性和底物专一性。DROSHA分子与同源二聚体结合 Dgcr8进行pri-miRNA加工，但也可能形成更高阶的复合体，因为 基因组中的pri-miRNAs促进了加工。我们最新的开创性低温电子显微镜(低温电子显微镜) EM)结构提供了微处理器-pri-miRNA复合体的原子模型。阐明了如何 蛋白质围绕RNA茎环组织，也揭示了每个远端(基端或顶端)是如何 独立识别，但也通过连接检测模块的分子尺相互连接。 拟议的研究建立在我们以前的成功基础上，因为结构框架将使我们能够 对如何实现大分子识别的新的基本见解。我们的总体目标是 从分子水平了解微处理器对pri-miRNAs的识别。我们假设核糖核酸 结构特征和上下文相关的序列首选项决定了每个个体的处理命运 PrI-miRNA。我们将剖析这种识别是如何在原基和顶端的每个连接处完成的。 MiRNAs。我们还将调查来自不同pri-miRNAs的不同RNA序列和结构如何影响 正确识别每个结合点，以揭示多部件机械的可塑性。我们之前的工作是 也让我们准备解决紧迫的问题，即pri-miRNAs的集群如何通过以下方式增强处理 微处理器，这对于我们对miRNA生物发生的全面理解至关重要，并具有深刻的 对解释过去和未来在各种领域取得的结果的影响 对miRNA基因的操纵。更好地掌握核心识别机制将有助于我们解释独特的 目标，如集群的pri-miRNAs。总而言之，拟议的研究将提供一个全面的 了解miRNAs的加工和调控，miRNAs是基因表达的重要调节因子。我们的工作是 破译结构如何影响RNA识别是一个根本上重要的问题，而且很可能 对于涉及miRNAs以外的结构化RNA的许多过程都很有洞察力。