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.

摘要 为了使亚细胞区域能够有效的特化和动态调节，许多细胞已经进化出局部的 然而，星形胶质细胞中这种翻译调节的基本原理是未知的。 长期以来，人们一直在神经元中研究，各种蛋白质的局部翻译被认为对突触至关重要， 影响学习和记忆的特殊变化。在本裁决的前一个周期，我们提供了证据， 星形胶质细胞还具有通过使用多种方法调节的局部翻译。在此，我们建议继续 这项工作，重点是关于这个迷人的新的基本生物现象的以下问题： 是决定何时以及哪些转录本在星形胶质细胞中局部翻译的调节语法？ 我们的中心模型是，转录本的非翻译区（UTR）中的元件负责它们的翻译。 富集或耗尽从核糖体在外周星形胶质细胞过程（PAP），通过UTR相互作用， RNA结合蛋白（RBP）和microRNA（miRNA）。然而，有数百种潜在的元素， 筛选，新的，可扩展的方法需要系统地表征RNA定位和翻译 在星形胶质细胞中，无论是在基线还是在对信号线索的响应中，都受到调节。此外，胶质形态学仅 在体内达到完全成熟，需要体内功能研究。因此，我们开发了一种新的方法， 突触神经小体-大规模平行报告基因分析（SN-MPRA），它允许我们评估功能效应 在体内同时检测数千个候选UTR元件。我们将应用此定义序列， 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 功能，我们将定义一个特定的RBP的作用，“震动”（QKI），在调节星形胶质细胞的本地翻译。 最后，为了理解如何以协调的方式调节转录本的集合，以便进行本地翻译， 我们将研究miRNA效应蛋白（Ago 2）沿着特定的miRNA在调节局部细胞凋亡中的作用。 在星形胶质细胞中的翻译。