Role of atlastin-1 in axonal development and degeneration of human neurons

描述(申请人提供)：轴突发育和退化受损与许多衰弱疾病有关，如遗传性痉挛截瘫(HSP)、肌萎缩侧索硬化症和周围神经病。HSP是由累及最长皮质脊髓束轴突的远端轴索病变引起的，导致肢体痉挛和无力。最常见的早发性HSP，SPG3A，是由atlastin-1基因突变引起的。这个基因编码atlastin-1蛋白，它是动力蛋白相关的大GTP酶超家族的成员。阿特拉斯汀-1对体外培养的大鼠皮质神经元的抑制作用 轴突的突起和延长。然而，atlastin-1活性改变如何导致HSP患者的轴突缺陷以及为什么特定的轴突退化在很大程度上尚不清楚。本研究的目的是建立SPG3A的人类神经元模型，以阐明HSP轴突缺陷的机制。本研究假设阿特拉斯汀基因突变选择性地导致皮质投射神经元轴突缺陷，这种作用主要是通过骨形态发生蛋白信号转导途径实现的。这一假说将通过以下两个目标得到验证：1)检测SPG3A患者和正常人(作为对照)产生的IPSCs来源的皮质PN中轴突的生长和运输；2)描述BMP信号在SPG3A轴突缺陷中的作用。通过比较对照组和SPG3A IPSCs来源的皮质三叉神经节、皮质中间神经元和脊髓运动神经元的轴突缺陷、atlastin-1活性和BMP信号变化，本研究将能够描绘HSP中特定细胞类型的缺陷及其潜在机制。通过敲除野生型(WT)神经元中的ATLASTIN-1和通过在SPG3A IPSCs中表达WT ATLASTIN-1，将证实ATLASTIN功能丧失与轴突表型之间的因果关系。此外，还将进行抢救性实验，以确定挽救轴突病理的潜在方法，如过度表达阿特拉斯汀或使用BMP拮抗剂治疗。综上所述，本研究将对atlastin-1和BMP信号在HSP病理中的作用提供有价值的见解，并为HSP轴突变性的抢救提供新的治疗方法。

DESCRIPTION (provided by applicant): Impaired axonal development and degeneration are implicated in many debilitating disorders, such as hereditary spastic paraplegia (HSP), amyotrophic lateral sclerosis, and periphery neuropathy. HSP is caused by distal axonopathy involving the longest corticospinal tract axons, leading to spasticity and weakness of the lower extremities. The most common early-onset form of HSP, SPG3A, is caused by mutations in the atlastin-1 gene. This gene encodes atlastin-1 protein, which is a member of the dynamin-related large GTPase superfamily. Knockdown of atlastin-1 in rat cortical neuron in vitro cultures inhibits the axonal outgrowth and elongation. However, how altered atlastin-1 activity leads to axonal defects and why specific axons degenerate in HSP patients are largely unclear. The goal of this proposed study is to establish human neuronal models of SPG3A to delineate the mechanisms underlying the axonal defects in HSP. This study's hypothesis is that atlastin mutations result in axonal defects selectively in cortical projection neurons (cortical PNs), and this effect is mediated mainly by dysregulated bone morphogenetic protein (BMP) signaling. This hypothesis will be tested by pursuing the following two aims: 1) to examine the axonal outgrowth and transport in cortical PNs derived from iPSCs that are generated from SPG3A patients and normal individuals (as controls); 2) to delineate the role of BMP signaling in the axonal defects in SPG3A. By comparing the axonal defects, atlastin-1 activity, and BMP signaling alterations in cortical PNs, cortical interneurons, and spinal motor neurons derived from control and SPG3A iPSCs, this study will be able to delineate the cell type-specific defects in HSP and the underlying mechanisms. The cause-effect relationship between loss of atlastin function and axonal phenotypes will be confirmed by knocking down atlastin-1 in wild-type (WT) neurons and by expressing WT atlastin-1 in SPG3A iPSCs. Moreover, rescue experiments will be performed to identify the potential approaches for rescuing the axonal pathology, such as overexpression of atlastin or treatment with BMP antagonists. Together, this study will provide valuable insights into the roles of atlastin-1 and BMP signaling in HSP pathology and developing new therapeutics for rescuing the axonal degeneration in HSP.