Defining the messenger RNP code in the brain

描述（由申请人提供）：RNA结合蛋白（RBP）与嵌入前信使RNA和成熟信使RNA中的功能性RNA元件相互作用，形成信使核糖核蛋白（mRNP）复合物。这些相互作用导致RNA加工事件的忠实执行，例如前mRNA选择性剪接，RNA稳定性和翻译控制。RBP和它们的RNA元件之间的相互作用的异常改变最终导致行为异常和神经发育缺陷，其通常可以表现为致命的疾病，如脊髓性肌萎缩症和肌萎缩性侧索硬化症或终身衰弱的行为异常，如Prader-Willi/Angelman Syndrome，精神分裂症和自闭症谱系障碍。这些发现强调了研究这些RBP在大脑中作用的重要性。特别是，我们的项目旨在系统地使用全基因组生物化学和生物信息学分析，识别小鼠大脑和人类神经元中RBP识别的功能性RNA元件。我们将开发一种新的人类多能干细胞资源，稳定表达可分化为人类神经元的标记RBP。这将使得能够使用尖端的基因组方法，如交联和免疫沉淀，然后进行高通量测序（CLIP-seq），以统一和系统的方式鉴定人神经RNA中50个RBP的RNA结合位点。为了揭示依赖于这些RBP直接结合的剪接、稳定性和翻译变化，我们将对mRNA（RNA-seq）和核糖体保护片段（RPF）进行高通量测序。最后，我们将利用我们的计算专业知识，使用大脑中的这种全基因组，多尺度，mRNP代码来构建预测模型。 公共卫生相关性：在过去的十年中，RNA结合蛋白（RBP）本身的缺陷或RBP的RNA结合位点之间的相互作用已被认为是神经系统疾病和精神障碍的原因。该建议旨在建立哺乳动物神经元中选定的RBP及其功能RNA元件之间的相互作用。这将是阐明大脑中这些RBP调控的RNA网络的关键的第一步。

DESCRIPTION (provided by applicant): RNA binding proteins (RBPs) interact with functional RNA elements embedded within pre- and mature messenger RNA to form messenger ribonucleoprotein (mRNP) complexes. These interactions result in the faithful execution of RNA processing events such as pre-mRNA alternative splicing, RNA stability and translational control. Aberrant alterations in the interactions between the RBPs and their RNA elements ultimately lead to behavioral abnormalities and neurological developmental defects, which can often manifest as fatal diseases such as Spinal Muscular Atrophy and Amyotrophic Lateral Sclerosis or life-long debilitating behavioral abnormalities such as Prader-Willi/Angelman Syndromes, Schizophrenia and Autism Spectrum Disorder. These findings underscore the importance of investigating the roles of these RBPs in the brain. In particular, our project is aimed at systematically, using genome-wide biochemical and bioinformatic assays, identifying the functional RNA elements that are recognized by RBPs in mouse brain and human neurons. We will develop a novel resource of human pluripotent stem cells stably expressing tagged RBPs that can be differentiated into human neurons. This will enable the identification of RNA binding sites of 50 RBPs in human neural RNAs in a uniform and systematic manner using cutting-edge genomic approaches such as cross-linking and immunoprecipitation followed by high-throughput sequencing (CLIP- seq). To reveal the splicing, stability, and translational changes that are dependent on direct binding of these RBPs we will perform high-throughput sequencing of mRNAs (RNA-seq) and ribosome-protected fragments (RPFs). Finally, we will leverage our computational expertise to build predictive models using this genome-wide, multi-scale, mRNP code in the brain. PUBLIC HEALTH RELEVANCE: Over the last decade, defects in RNA binding proteins (RBPs) themselves or the interactions between RNA binding sites of RBPs have become recognized as increasingly responsible for neurological disease and mental disorders. This proposal seeks to establish the interactions among selected RBPs and their functional RNA elements in mammalian neurons. This will be a crucial first step in elucidating the RNA networks regulated by these RBPs in the brain.