Mechanism and Function Of MBNL Mediated mRNA Localization in Neuronal Development and Neurologic Disease

MBNL介导的mRNA定位在神经元发育和神经系统疾病中的机制和功能

• 批准号: 10553695
• 负责人: GARY J BASSELL
• 金额: $ 41.2万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10553695
• 关键词: Adult; Alternative Splicing; Axon; Basic Science; Biochemical; Brain Diseases; CUG repeat; Cell Nucleus; Cell physiology; Cells; Cellular biology; Charcot-Marie-Tooth Disease; Complex; Cytoplasm; Cytoplasmic Granules; Cytoskeleton; Dendrites; Destinations; Development; Disease; Distal; Elements; Exhibits; FMRP; Fragile X Syndrome; Genomics; Hippocampus; Impairment; In Vitro; Investigation; Kinesin; Kinetics; Link; Maintenance; Mediating; Membrane; Messenger RNA; Microtubules; Mitochondria; Modeling; Morphology; Motor; Mus; Muscle; Mutate; Myotonic Dystrophy; Nerve Degeneration; Neurites; Neurologic; Neurons; Neuropil; Pathogenesis; Patients; Process; Property; Proteins; RNA; RNA Splicing; RNA Transport; RNA-Binding Proteins; Records; Regulation; Reporting; Research; Resolution; Signal Transduction; Site; Skeletal Muscle; Symptoms; Synapses; Synaptic Vesicles; Synaptic plasticity; Translating; Translations; Variant; biophysical properties; cell motility; live cell imaging; mouse model; nervous system disorder; neuron development; novel; postsynaptic; proteostasis; synaptic function; vesicular release

The development and maintenance of neurons and synaptic connections are highly complex processes, in part due to the massive cytoplasmic volume and complex branching morphology of axons and dendrites. As one mechanism, it is well appreciated that RNA localization and local translation are required to precisely regulate protein homeostasis at synapses. Indeed, loss of FMRP in Fragile X Syndrome, or other impairments to RNA localization and local translation at synapses, likely contribute to brain disorders. To better understand RNA localization and local translation in neurons, we must elucidate the RNA cis-elements, RBP trans-factors, and cytoskeletal motors mediating these processes. Although ongoing efforts have demonstrated how RNA binding proteins (RBPs) can regulate local translation at post-synaptic sites, there still exists a major gap in our understanding of how RBPs transport RNAs to regulate synaptic function. Fortunately, recent observations provide clues about fruitful lines of investigation. For example, multiple studies report that distally localized RNAs are enriched for cis-elements targeted by Muscleblind-like (MBNL) proteins. Although these observations suggest that MBNL may be a major player in localizing RNAs to the pre- and post-synapse, we still lack a mechanistic understanding for how MBNL proteins may achieve this task, or what functions depend on MBNL-mediated RNA localization. This line of research has important implications for the neurological disease myotonic dystrophy (dystrophia myotonica, DM), in which MBNLs are depleted by toxic CUG repeats. Therefore, an emerging hypothesis is that RNA localization functions of MBNL are important for proper synapse function, and that mis-localized RNAs might account for some neurological features of DM patients, particularly early in disease. Here, using MBNL depletion and DM-associated models, we propose to identify specific functions for the localization of MBNL targets. Aim 1 will elucidate mechanisms of MBNL-mediated mRNA localization in neurons. We will define the RNA targets that are localized by MBNL in the pre- and post-synapse. We will characterize dynamic properties of motile MBNL RNA granules in live neurons and identify cytoskeletal motors and adaptors associated with these granules. Using genomics, live cell imaging, and biochemical approaches, we will establish mechanisms of how MBNL-interacting RNAs are transported. Aim 2 will define functions conferred by MBNL-dependent RNA localization using models of synapse development and function, and models of myotonic dystrophy. By depleting cytoplasmic MBNL and other proteins required for MBNL-dependent RNA localization, we will assess cellular functions dependent on this process. We will identify specific neuronal functions, such as synaptic vesicle release, that depend on proper localization of mRNAs by MBNL proteins. The impact of this research is to better understand how RNA localization and local translation confers important synaptic functions and how they may go awry in DM. As few RNA binding proteins have been linked to motors, this may evolve into a unifying model for mRNA transport to synapses.

神经元和突触连接的发展和维持是高度复杂的过程，在一定程度上 由于轴突和树突的大量胞质体积和复杂的分支形态。作为一个整体 机制，人们很好地理解，需要RNA本地化和本地翻译来精确调控 突触的蛋白质动态平衡。事实上，脆性X综合征中FMRP的丢失，或RNA的其他损害 突触的定位和局部翻译可能会导致大脑紊乱。为了更好地理解RNA 在神经元的定位和局部翻译方面，我们必须阐明RNA顺式元件、RBP反式因子和 细胞骨架马达调节这些过程。尽管正在进行的研究已经证明了RNA结合 蛋白质(RBPs)可以调节突触后部位的局部翻译，但在我们的 了解限制性商业惯例如何运输RNA来调节突触功能。幸运的是，最近的观察 提供富有成效的调查线索。例如，多项研究报告称，远端定位 RNA富含类似蝇毒样蛋白(MBNL)靶向的顺式元件。尽管这些 观察表明，MBNL可能是将RNA定位到突触前和突触后的主要参与者 仍然缺乏对MBNL蛋白如何完成这一任务或依赖于哪些功能的机械性理解 关于MBNL介导的RNA定位。这一研究路线对神经学具有重要的意义 病理性强直性肌营养不良(dystrophia myotonica，DM)，其中MBNL被有毒的CUG重复序列耗尽。 因此，一个新的假设是，MBNL的RNA定位功能对于正常的 突触功能，错误定位的RNA可能解释了糖尿病患者的一些神经特征， 尤其是在疾病的早期。在这里，使用MBNL耗尽和DM相关模型，我们建议识别 MBNL目标本地化的具体功能。目标1将阐明MBNL介导的机制 信使核糖核酸在神经元的定位。我们将定义MBNL在突触前和突触后定位的RNA靶标。我们将表征活神经元中可移动的MBNL RNA颗粒的动态特性并识别 与这些颗粒相关的细胞骨架马达和适配器。使用基因组学、活细胞成像和 通过生化方法，我们将建立MBNL相互作用的RNA如何运输的机制。目标2 将使用突触发育模型定义依赖MBNL的RNA本地化所赋予的功能 和功能，以及强直性肌营养不良模型。通过耗尽细胞质MBNL和其他所需的蛋白质 对于MBNL依赖的RNA定位，我们将评估依赖于这一过程的细胞功能。我们会 确定特定的神经元功能，如突触小泡释放，这取决于适当的定位 MBNL蛋白表达的mRNAs。这项研究的影响是更好地了解RNA本地化和局部化 翻译赋予了重要的突触功能，以及它们在DM中可能出现的错误。作为很少的RNA结合 蛋白质已经与马达相连，这可能进化成一个统一的模型，将信使核糖核酸运输到突触。