Survival Motor Neuron (SMN) function in motoneuron development

项目摘要 了解运动神经元功能障碍的机制基础及其在运动神经元疾病中的作用将填补 神经科学的重大差距和推进治疗肌营养不良等破坏性疾病的新方法 侧索硬化症(ALS)、遗传性运动神经病和脊肌萎缩症(SMA)。这些疾病困扰着人们。 在美国，每年有超过10万的成年人、婴儿和儿童。ALS和SMA特别 破坏性疾病导致瘫痪和死亡，通常在确诊后的几年内。这些基因的遗传学 疾病表明，运动神经元特别容易受到负责关键rna的蛋白质的缺陷的影响。 处理功能。然而，运动神经元容易受到RNA加工缺陷的影响的确切原因并不是 明白了。这个项目的科学基础是机械地阐明如何错误地处理 RNA可以破坏运动神经元的功能，导致运动神经元死亡。阐明运动神经元的特异性 由这些突变引起的RNA加工缺陷对于理解这两种正常的运动神经元是必不可少的 和疾病，并将指导急需的治疗方法。为了解决这个问题，我们专注于 广泛表达存活运动神经元(SMN)蛋白和运动神经元疾病SMA。SMA是一种 运动神经元疾病，影响婴儿/儿童，由低存活运动神经元(SMN)蛋白水平引起。 SMN在RNA代谢的许多方面起着重要作用。然而，SMN的关键RNA处理功能在 运动神经元尚未解决。证据支持SMN与多种神经元RNA结合蛋白相互作用 (限制性商业惯例)在发育过程中稳定RNA和/或将RNA运输到轴突和树突。使用独特的斑马鱼 我们已经建立的模型表明，SMN是正常脊椎动物运动神经元所必需的 发育包括树突形成和运动轴突长出和树枝形成。这是一项关键发现， 提示SMA不是退行性病变，但运动神经元功能障碍是运动神经元功能低下所致。 发育导致神经元衰竭。我们假设SMN与神经元RBPs和它们的 以发育调节的方式引导运动神经元发育的货物RNA，包括轴突 向外生长和分枝，树突形成和突触形成。为了测试这一点，我们将回答三个问题 基本问题：在运动神经元发育过程中，SMN：RBP复合体是什么？这些限制性商业惯例中的缺陷是如何 会影响运动神经元发育吗？这些复合体中有哪些RNA，当SMN或 限制性商业惯例缺失还是减少了？我们的所有实验都将在活体运动神经元中进行，相关的 细胞类型，并使用广泛的实验方法，如生物化学，质量分光光度， RNAseq、单神经元成像和遗传学。这些实验的数据将对 了解RNA参与正常运动神经元发育、SMA和其他运动神经元疾病，如 作为肌萎缩侧索硬化。此外，我们的方法将严格测试SMN：RBP复合体及其相关的重要性 揭示运动神经元生物学基本分子机制的RNA。

Project Summary Understanding the mechanistic basis of motoneuron dysfunction and its role in motoneuron diseases would fill a major gap in neuroscience and advance new approaches for treating devastating diseases such as amyotrophic lateral sclerosis (ALS), hereditary motor neuropathy and spinal muscular atrophy (SMA). These diseases afflict over one hundred thousand adults, infants, and children per year in the US. ALS and SMA are particularly devastating diseases resulting in paralysis and death often within a few years of diagnosis. The genetics of these diseases indicates that motoneurons are particularly vulnerable to defects in proteins tasked with critical RNA processing functions. However, exactly why motoneurons are vulnerable to RNA processing defects is not understood. The scientific rationale for this project is to elucidate mechanistically how mishanding of RNAs can disrupt motoneuron function and lead to motoneuron death. Elucidating the motoneuron-specific RNA processing defects caused by these mutations is essential for understanding motoneurons in both normal and diseased conditions and will direct critically needed therapeutics. To tackle this issue, we focus on the ubiquitously expressed survival motor neuron (SMN) protein and the motoneuron disease SMA. SMA is a motoneuron disease that affects infants/children and is caused by low survival motor neuron (SMN) protein levels. SMN functions in many aspects of RNA metabolism. However, the critical RNA handling function of SMN in motoneurons is unresolved. Evidence supports that SMN interacts with various neuronal RNA binding proteins (RBPs) that stabilize and/or transport RNAs to axons and dendrites during development. Using unique zebrafish models that we have generated, we have shown that SMN is required for normal vertebrate motoneuron development including dendrite formation and motor axon outgrowth and arborization. This is a key finding and reveals that SMA is not a degenerative defect, but the motoneuron dysfunction is caused by poor motoneuron development leading to neuronal failure. We hypothesize that SMN associates with neuronal RBPs and their cargo RNAs in a developmentally regulated manner to direct motoneuron development including axon out growth and branching, dendrite formation, and synapse formation. To test this we will answer three essential questions: What SMN:RBP complexes are in developing motoneurons? How do defects in these RBPs affect motoneuron development? What RNAs are in these complexes, and how are they affected when SMN or the RBPs are missing or decreased? All of our experiments will be performed in vivo in motoneurons, the relevant cell type and use a broad range of experimental approaches such as biochemistry, mass spectrophotometry, RNAseq, single neuron imaging and genetics. Data from these experiments will have broad implications for understanding RNA involvement in normal motoneuron development, SMA, and other motoneuron diseases such as ALS. In addition, our approach will rigorously test the importance of SMN:RBP complexes and their associated RNAs revealing a fundamental molecular mechanism in motoneuron biology.