A Neural Development mRNA Decay Network

神经发育 mRNA 衰减网络

• 批准号: 8560494
• 负责人: Michael Cleary
• 金额: $ 26.99万
• 依托单位: UNIVERSITY OF CALIFORNIA, MERCED
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8560494
• 关键词: Affect; Affinity Chromatography; Beryllium; Binding; Binding Proteins; Biochemical; Biological Assay; Cells; Confocal Microscopy; Congenital Abnormality; Coupled; Data; Defect; Detection; Development; Disease; Drosophila genus; Electrophoretic Mobility Shift Assay; Elements; Embryo; Embryonic Development; Embryonic Nervous System; Foundations; Fragile X Syndrome; Gene Expression; Generations; Genes; Genetic; Genetic Transcription; Human; Image; Immunoprecipitation; Impairment; In Vitro; Indium; Individual; Infant Mortality; Inherited; Kinetics; Maps; Mass Spectrum Analysis; Measurement; Measures; Mediating; Messenger RNA; Methods; MicroRNAs; Mitotic; Modeling; Molecular Genetics; Mutation; Nervous system structure; Neurologic; Neurons; Pathway interactions; Population; Porifera; Proliferating; Psyche structure; RNA; RNA Decay; RNA Interference; RNA Sequences; RNA Stability; RNA-Binding Proteins; RNA-Protein Interaction; Regulation; Regulatory Element; Reporter; Resolution; Sampling; Site; Spinal Muscular Atrophy; Stem cells; Structure; System; Techniques; Testing; Transcript; Work; base; candidate identification; cell type; embryo cell; genome-wide; genome-wide analysis; in vivo; insertion/deletion mutation; mutant; nervous system development; network models; neuroblast; neurodevelopment; null mutation; programs; public health relevance; recombinant RNA; relating to nervous system

DESCRIPTION (provided by applicant): Gene expression during development is often exclusively studied at the level of transcription. However, an additional and essential level of control occurs via post-transcriptional mRNA decay. Regulation of mRNA decay is particularly important during nervous system development, where the structure of neurons requires selective stabilization of mRNAs far from their site of synthesis and the generation of cellular diversity requires the programmed decay of mRNAs that regulate proliferation and differentiation. Defective mRNA decay has been implicated in many neurological birth defects, including fragile X-syndrome (the most common form of hereditary mental impairment) and spinal muscular atrophy (a leading genetic cause of infant mortality). The study of mRNA decay during embryonic development has previously been hindered by the lack of methods allowing in vivo, cell type-specific measurements of transcript stability. We have developed a technique (based on TU-tagging methods) that overcomes this technical challenge and allows neural-specific, genome-wide measurements of mRNA decay in intact Drosophila embryos. This technique provides the foundation for a systems-level approach that will allow us to construct a Neural Development mRNA Decay Network. This network will contain the following information: the decay rates of all mRNAs expressed in the embryonic nervous system, the cis-elements that target these mRNAs for decay, the trans-acting RNA-binding proteins and miRNAs that bind these cis-elements, and the spatial dynamics of mRNA - RNA-binding protein interactions. This information will be obtained using a systems-level approach that combines genome-wide analysis of mRNA decay rates in wildtype and RBP mutant embryos, computational identification of candidate linear and structural cis-elements, molecular genetic definition of cis element effects on transcript stability in vivo, biochemical analysis of RBP-mRNA interactions in vitro and in vivo, and imaging of fluorescent mRNA - RBP interactions within neurons. This work will generate a comprehensive and predictive network map of neural mRNA decay dynamics, thus filling a significant gap in current models of gene expression during neural development.

描述（由申请人提供）：发育过程中的基因表达通常仅在转录水平上进行研究。然而，额外的和必要的控制水平通过转录后mRNA衰变发生。mRNA衰变的调节在神经系统发育期间特别重要，其中神经元的结构需要远离其合成位点的mRNA的选择性稳定，并且细胞多样性的产生需要调节增殖和分化的mRNA的程序性衰变。缺陷性mRNA衰变与许多神经系统出生缺陷有关，包括脆性X综合征（遗传性精神障碍的最常见形式）和脊髓性肌萎缩症（婴儿死亡的主要遗传原因）。胚胎发育过程中mRNA衰变的研究以前一直受到阻碍，缺乏方法，允许在体内，细胞类型特异性的转录稳定性测量。我们已经开发了一种技术（基于TU标记方法），克服了这一技术挑战，并允许神经特异性，全基因组测量完整果蝇胚胎中的mRNA衰变。这项技术为系统级方法提供了基础，使我们能够构建神经发育mRNA衰变网络。该网络将包含以下信息：胚胎神经系统中表达的所有mRNA的衰变速率、靶向这些mRNA衰变的顺式元件、反式作用RNA结合蛋白和结合这些顺式元件的miRNA以及mRNA -RNA结合蛋白相互作用的空间动力学。该信息将使用系统水平的方法获得，该方法结合了野生型和RBP突变胚胎中mRNA衰减率的全基因组分析、候选线性和结构顺式元件的计算鉴定、顺式元件的分子遗传学定义、顺式元件的分子遗传学定义、顺式元件的分子遗传学定义和顺式元件的分子遗传学定义。 元件对体内转录物稳定性的影响，体外和体内RBP-mRNA相互作用的生物化学分析，以及神经元内荧光mRNA - RBP相互作用的成像。这项工作将产生一个全面的和预测性的神经mRNA衰减动力学网络图，从而填补了目前神经发育过程中基因表达模型的重大空白。