Cold Spring Harbor Laboratory, 1 Bungtown Rd, Cold Spring Harbor, NY 11724, USA; Graduate Program in Neuroscience, Life Sciences 550, SUNY at Stony Brook, Stony Brook, NY 11794, USA; Dart Neuroscience LLC, 12278 Scripps Summit Dr., San Diego, CA 92131, USA.
Cold Spring Harbor Laboratory, 1 Bungtown Rd, Cold Spring Harbor, NY 11724, USA; Dart Neuroscience LLC, 12278 Scripps Summit Dr., San Diego, CA 92131, USA.
Neurobiol Dis. 2018 Jun;114:153-163. doi: 10.1016/j.nbd.2018.03.003. Epub 2018 Mar 7.
Parkinson's disease (PD) is a progressive motor neurodegenerative disorder, characterized by a selective loss of dopaminergic neurons in the substantia nigra. The complexity of disease etiology includes both genetic and environmental factors. No effective drug that can modify disease progression and protect dopamine neurons from degeneration is presently available. Human α-Synuclein A30P (A30P) is a mutant gene identified in early onset PD and showed to result selective dopamine neuron loss in transgenic A30P flies and mice. Paraquat (PQ) is an herbicide and an oxidative stress generator, linked to sporadic PD. We hypothesized that vital PD modifier genes are conserved across species and would show unique transcriptional changes to oxidative stress in animals expressing a PD-associated gene, such as A30P. We also hypothesized that manipulation of PD modifier genes would provide neuroprotection across species. To identify disease modifier genes, we performed two independently-duplicated experiments of microarray, capturing genome-wide transcriptional changes in A30P flies, chronically fed with PQ-contaminated food. We hypothesized that the best time point of identifying a disease modifier gene is at time when flies showed maximal combined toxicity of A30P transgene and PQ treatment during an early stage of disease and that effective disease modifiers gene are those showing transcriptional changes to oxidative stress in A30P expressing and not in wild type animals. Fly Neprilysin3 (Nep3) is one identified gene that is highly conserved. Its mouse and human homolog is endothelin-converting enzyme-1 (Ece1). To investigate the neuroprotective effect of Ece1, we used NS1 cells and mouse midbrain neurons expressing A30P, treated with or without PQ. We found that ECE1 expression protected against A30P toxicity on cell viability, on neurite outgrowth and ameliorated A30P accumulation in vitro. Expression of ECE1 in vivo suppressed dopamine neuron loss and alleviated the corresponding motor deficits in mice with A30P-expression. Our study leverages a new approach to identify disease modifier genes using a stress-enhanced PD animal model.
帕金森病（PD）是一种进行性运动神经退行性疾病，其特征是黑质中多巴胺能神经元的选择性丧失。疾病病因的复杂性包括遗传和环境因素。目前尚无有效的药物可以改变疾病进展并防止多巴胺神经元变性。人类α-突触核蛋白 A30P（A30P）是在早发性 PD 中鉴定的突变基因，并在转基因 A30P 果蝇和小鼠中显示出选择性多巴胺神经元丢失。百草枯（PQ）是一种除草剂和氧化应激生成剂，与散发性 PD 有关。我们假设重要的 PD 修饰基因在物种间是保守的，并且在表达与 PD 相关基因（如 A30P）的动物中对氧化应激会显示出独特的转录变化。我们还假设 PD 修饰基因的操作将在物种间提供神经保护。为了鉴定疾病修饰基因，我们进行了两次独立复制的微阵列实验，以捕获 A30P 果蝇的全基因组转录变化，这些果蝇长期食用被 PQ 污染的食物。我们假设鉴定疾病修饰基因的最佳时间点是在 A30P 转基因和 PQ 处理的最大联合毒性显示在疾病的早期阶段，并且有效的疾病修饰基因是那些在 A30P 表达时对氧化应激显示转录变化而不是在野生型动物中显示的基因。果蝇神经肽酶 3（Nep3）是一个高度保守的已鉴定基因。其小鼠和人类同源物是内皮素转换酶 1（Ece1）。为了研究 Ece1 的神经保护作用，我们使用 NS1 细胞和表达 A30P 的小鼠中脑神经元，用或不用 PQ 处理。我们发现 ECE1 的表达可防止 A30P 毒性对细胞活力、神经突生长的影响，并在体外改善 A30P 的积累。ECE1 的体内表达可抑制 A30P 表达的小鼠中多巴胺神经元的丢失，并减轻相应的运动缺陷。我们的研究利用了一种新方法，使用增强应激的 PD 动物模型来鉴定疾病修饰基因。
