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: 10334425
• 负责人: GARY J BASSELL
• 金额: $ 42.12万
• 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/10334425
• 关键词: Adult; Alternative Splicing; Axon; Basic Science; Biochemical; Biophysics; Brain Diseases; CUG repeat; Cell Nucleus; Cell physiology; Cells; Cellular biology; Charcot-Marie-Tooth Disease; Complex; Cytoplasm; Cytoplasmic Granules; Dendrites; Destinations; Development; Disease; Distal; Elements; Exhibits; FMRP; Fragile X Syndrome; Genomics; Hippocampus (Brain); 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; Symptoms; Synapses; Synaptic Vesicles; Synaptic plasticity; Translating; Translations; Variant; base; cell motility; live cell imaging; mouse model; nervous system disorder; neuron development; novel; proteostasis; skeletal muscle wasting; 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育和维持是高度复杂的过程，部分地， 这是由于轴突和树突的大量细胞质体积和复杂的分支形态。作为一个 由于RNA的定位和局部翻译是精确调控细胞内RNA表达的重要机制， 突触的蛋白质稳态事实上，脆性X综合征中FMRP的缺失或其他RNA损伤 突触的局部化和局部翻译，可能会导致大脑疾病。为了更好地理解RNA 定位和局部翻译的神经元，我们必须阐明RNA顺式元件，RBP反式因子， 细胞骨架马达介导这些过程。尽管不断的努力已经证明了RNA结合 尽管RBP蛋白（RBP）可以调节突触后位点的局部翻译，但我们的研究仍存在重大空白。 了解RBP如何转运RNA来调节突触功能。幸运的是，最近的观察 为卓有成效的调查提供线索例如，多项研究报告称， RNA富含肌盲样（MBNL）蛋白靶向的顺式元件。虽然这些 观察表明，MBNL可能是一个主要的球员在本地化RNA的前和后突触，我们 仍然缺乏对MBNL蛋白如何实现这一任务的机制理解，或者什么功能依赖于 MBNL介导的RNA定位。这一系列的研究对神经学有着重要的意义。 疾病强直性肌营养不良（强直性肌营养不良，DM），其中MBNL被毒性CUG重复耗尽。 因此，一个新兴的假设是，MBNL的RNA定位功能对于适当的 突触功能，错误定位RNA可能解释了DM患者的一些神经学特征， 尤其是在疾病早期。在这里，使用MBNL耗尽和DM相关模型，我们建议识别 用于MBNL靶标定位的特定功能。目的1将阐明MBNL介导的 神经元中的mRNA定位。我们将定义MBNL在突触前和突触后定位的RNA靶点。我们将描述活神经元中运动的MBNL RNA颗粒的动力学特性， 与这些颗粒相关的细胞骨架马达和适配器。利用基因组学，活细胞成像， 通过生物化学方法，我们将建立MBNL相互作用RNA如何转运的机制。目的2 将使用突触发育模型定义MBNL依赖的RNA定位所赋予的功能 和功能，以及强直性肌营养不良的模型。通过消耗细胞质MBNL和其他所需的蛋白质， 对于MBNL依赖的RNA定位，我们将评估依赖于该过程的细胞功能。我们将 确定特定的神经元功能，如突触囊泡释放，这取决于适当的定位， MBNL蛋白的mRNA。这项研究的影响是更好地了解RNA定位和局部 翻译赋予重要的突触功能，以及它们在DM中如何出错。很少RNA结合 蛋白质已经连接到马达，这可能会演变成一个统一的模型mRNA运输到突触。