Regulation of local translation in glia

神经胶质细胞局部翻译的调控

• 批准号: 10737355
• 负责人: JOSEPH D DOUGHERTY
• 金额: $ 54.99万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10737355
• 关键词: Astrocytes; Award; Belief; Binding; Binding Sites; Biological Assay; Biological Phenomena; Biological Process; Brain; Cell Nucleus; Cells; Complex; Coupling; Cues; Databases; Dendritic Spines; Elements; Face; Family; Fluorescent in Situ Hybridization; Gene Expression; Genes; Image; Individual; Learning; Machine Learning; Mediating; Memory; Messenger RNA; Methods; MicroRNAs; Minority; Modeling; Molecular; Morphology; Mutation; Neurodegenerative Disorders; Neuroglia; Neurons; Pathway interactions; Peripheral; Phosphorylation; Phosphotransferases; Physiological; Post-Translational Protein Processing; Process; Proteins; Proteome; RNA; RNA-Binding Proteins; Regulation; Reporter; Response Elements; Ribosomes; Role; Signal Transduction; Site; Slice; Specificity; Structure; Supporting Cell; Synapses; Synaptosomes; Testing; Trans-Activators; Transcript; Transcriptional Regulation; Translating; Translation Process; Translational Regulation; Translational Repression; Translations; Untranslated Regions; Validation; Work; cell type; density; design; experimental study; fascinate; in vivo; inhibitor; mRNA Translation; overexpression; pharmacologic; response; tool; translatome

ABSTRACT To enable efficient specialization and dynamic regulation of subcellular regions, many cells have evolved local translation of mRNA - yet the fundamental principles of such translation regulation in astrocytes are unknown. Long studied in neurons, local translation of a variety of proteins is thought to be essential for the synapse- specific changes that underlay learning and memory. In the prior cycle of this award we provided evidence astrocytes also have a regulated local translation by using a variety of approaches. Here, we propose to continue this work, focusing on the following question regarding this fascinating new basic biological phenomenon: what is the regulatory grammar that determines when and which transcripts are locally translated in astrocytes? Our central model is that elements in the untranslated regions (UTRs) of transcripts are responsible for their enrichment or depletion from ribosomes in peripheral astrocyte processes (PAPs), via UTR interactions with RNA binding proteins (RBPs) and microRNAs (miRNAs). However, with hundreds of potential elements to screen, new, scalable methods are needed to systematically characterize how RNA localization and translation is regulated in astrocytes, both at baseline and in response to signaling cues. Furthermore, glial morphology only reaches full maturity in vivo, requiring in vivo functional studies. Therefore, we have developed a new method, a synaptoneurosome–massively parallel reporter assay (SN-MPRA) which allows us to assess functional effects of thousands of candidate UTR elements in vivo simultaneously. We will apply this to define the sequences that modulate RNA localization in astrocytes. Furthermore, to better understand how a subset of these elements function, we will define the role of a specific RBP, ‘quaking’ (QKI), in modulating local translation in astrocytes. Finally, to understand how sets of transcripts might be regulated in a coordinated fashion for local translation, we will examine the role of miRNA effector proteins (Ago2) along with specific miRNAs in regulating local translation in astrocytes.

抽象的 为了实现亚细胞区域的有效专业化和动态调节，许多细胞已经进化出局部 mRNA 的翻译 - 然而星形胶质细胞中这种翻译调节的基本原理尚不清楚。 在神经元中进行了长期研究，多种蛋白质的局部翻译被认为对于突触至关重要。 学习和记忆基础上的具体变化。在该奖项的前一个周期中，我们提供了证据 星形胶质细胞还通过使用多种方法进行受调节的局部翻译。在此，我们建议继续 这项工作，重点关注以下关于这一令人着迷的新基本生物现象的问题：什么 决定何时以及哪些转录物在星形胶质细胞中本地翻译的调节语法是什么？ 我们的中心模型是转录本非翻译区 (UTR) 中的元素负责其 通过 UTR 相互作用，外周星形胶质细胞过程 (PAP) 中核糖体的富集或消耗 RNA 结合蛋白 (RBP) 和 microRNA (miRNA)。然而，有数百种潜在元素 筛选，需要新的、可扩展的方法来系统地表征 RNA 定位和翻译的方式 在星形胶质细胞中，无论是在基线还是响应信号线索，都受到调节。此外，仅神经胶质形态 在体内达到完全成熟，需要进行体内功能研究。因此，我们开发了一种新方法， 突触神经体-大规模并行报告基因测定（SN-MPRA）使我们能够评估功能效应 同时体内数千个候选UTR元件。我们将应用它来定义序列 调节星形胶质细胞中的 RNA 定位。此外，为了更好地理解这些元素的子集如何 函数中，我们将定义特定 RBP“颤动”（QKI）在调节星形胶质细胞局部翻译中的作用。 最后，为了了解如何以协调的方式调节转录本集以进行本地翻译， 我们将研究 miRNA 效应蛋白 (Ago2) 以及特定 miRNA 在调节局部 星形胶质细胞中的翻译。