Shedding light on the role of RNA binding protein-mediated RNA regulation in synaptic plasticity

揭示 RNA 结合蛋白介导的 RNA 调节在突触可塑性中的作用

• 批准号: 10285142
• 负责人: Ruth A Singer
• 金额: $ 6.64万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2022-12-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10285142
• 关键词: Affinity Chromatography; Alleles; Alternative Splicing; Axon; Bioinformatics; Biological Assay; Biology; Brain; Breeding; Caliber; Cell Nucleus; Cells; Complex; DNA; Data; Data Set; Dendrites; Disease; Environment; FMR1; Fluorescent in Situ Hybridization; Foundations; Gene Expression; Genes; Genetic Transcription; High-Throughput Nucleotide Sequencing; Hippocampus (Brain); Immediate-Early Genes; Immunoprecipitation; Knowledge; Laboratories; Learning; Light; Lighting; Measures; Mediating; Memory; Messenger RNA; Methodology; Modeling; Molecular; Mus; Neurologic; Neurons; Neuropil; Neurosciences; Optics; Pathologic; Poly A; Population; Process; Protein Biosynthesis; Proteome; RNA; RNA Binding; RNA metabolism; RNA-Binding Proteins; Regulation; Research; Resolution; Rest; Role; Site; Structure; Synapses; Synaptic plasticity; Technology; Testing; Time; Transcript; Translations; Universities; Up-Regulation; calmodulin-dependent protein kinase II; cell type; crosslink; excitatory neuron; experience; experimental study; genetic information; in vivo; insight; interest; mRNA Precursor; mouse model; nervous system disorder; neuronal cell body; novel; novel strategies; optogenetics; protein distribution; relating to nervous system; response; unpublished works

Project Summary Neurons have highly specialized structures and functions; their genetic information is often located a great distance from the sites where information gets transmitted, and they must dynamically alter the synaptic proteome in response to neural activity. These challenges indicate that neurons have evolved unique regulatory mechanisms to meet functional demands. The uniformity of DNA across different cell types suggests that how genetic information is unfolded and processed underlies cellular diversity. In this view, careful examination of the regulation of RNA metabolism, how pre-mRNA gene copies are alternatively spliced and polyadenylated, edited, localized, and translationally regulated, will offer a new avenue towards understanding complex processes, such as synaptic plasticity underlying learning and memory. This notion has driven molecular neuroscientists to search for factors that localize and regulate synaptic RNAs. We are only beginning to compile a list of these key regulators, such as RNA binding proteins (RBPs), particularly in the synapses of hippocampal neurons that are involved in memory, and to date very little is known about how these factors regulate RNA. Our laboratory has recently generated a new platform for cell-specific Crosslinked Immunoprecipitation (CLIP) of RBPs in the living brain of mice (cTag-CLIP), which has furthered our understanding of the cell-specific regulatory functions of RBPs; however, this technology is limited to looking at steady state RNA regulation. Here we described a novel approach to uncover the role of neuronal RBP-mediated RNA regulation in the context of synaptic plasticity. This methodology, termed opto-CLIP, will combine the cell type-specific resolution afforded by cTag-CLIP with the unprecedented precision of optogenetics to achieve non-invasive optical control of specific neurons. Once established, we will increase the cellular resolution by performing opto-CLIP in distinct subcellular compartments to assess local RNA regulation associated with neuronal function. This study will further our understanding of RBP-mediated RNA regulation, enhance our knowledge of the role of RNA metabolism in synaptic plasticity, and provide new insight into the pathological mechanisms underlying neurological disorders. In conclusion, I am confident that my experience studying RNA biology, the Darnell lab's foundation in neuroscience and CLIP, and the rich research environment at Rockefeller University will help me succeed in using optogenetics to study the role of RBP-mediated RNA regulation in synaptic plasticity.

项目摘要 神经元具有高度专业化的结构和功能;它们的遗传信息通常位于很大的位置。 它们必须动态地改变突触， 蛋白质组对神经活动的反应。这些挑战表明，神经元已经进化出独特的调节机制， 满足功能需求。不同细胞类型的DNA的一致性表明， 遗传信息的展开和处理是细胞多样性的基础。在这种情况下，仔细检查 RNA代谢的调节，前mRNA基因拷贝如何被选择性剪接和多腺苷酸化，编辑， 本地化和规范化，将提供一个新的途径来理解复杂的过程， 作为学习和记忆基础的突触可塑性。这一概念促使分子神经科学家们寻找 定位和调节突触RNA的因子。我们才刚刚开始编制这些关键要素的清单 调节剂，如RNA结合蛋白（RBP），特别是在海马神经元的突触中， 这些因子与记忆有关，迄今为止，人们对这些因子如何调节RNA知之甚少。本实验室 最近产生了一个新的平台，细胞特异性交联免疫沉淀（CLIP）的RBP在生活中 小鼠脑（cTag-CLIP），这进一步加深了我们对细胞特异性调节功能的理解， 然而，这项技术仅限于观察稳态RNA调控。这里我们描述了一部小说 揭示神经元RBP介导的RNA调节在突触可塑性中的作用的方法。这 一种称为opto-CLIP的方法将联合收割机结合cTag-CLIP提供的细胞类型特异性分辨率与 光遗传学前所未有的精确度，以实现特定神经元的非侵入性光学控制。一旦 我们将通过在不同的亚细胞区室中进行opto-CLIP来提高细胞分辨率 以评估与神经元功能相关的局部RNA调节。这项研究将进一步加深我们对 RBP介导的RNA调节，增强了我们对RNA代谢在突触可塑性中的作用的认识， 为神经系统疾病的病理机制提供了新的见解。总之，我 我相信，我学习RNA生物学的经验，达内尔实验室在神经科学和CLIP方面的基础， 洛克菲勒大学丰富的研究环境将帮助我成功地利用光遗传学来研究 RBP介导的RNA调节在突触可塑性中的作用。