A De Novo RAPGEF2 Variant Identified in a Sporadic Amyotrophic Lateral Sclerosis Patient Impairs Microtubule Stability and Axonal Mitochondria Distribution
A De Novo RAPGEF2 Variant Identified in a Sporadic Amyotrophic Lateral Sclerosis Patient Impairs Microtubule Stability and Axonal Mitochondria Distribution
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DOI：
10.5607/en.2018.27.6.550
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发表时间：
2018-12-01
影响因子：
2.4
通讯作者：
Lee, Seungbok
中科院分区：
文献类型：
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作者：
Heo, Keunjung;Lim, Su Min;Lee, Seungbok
肌萎缩侧索硬化症 (ALS) 是一种致命的神经退行性疾病，通常与微管异常和线粒体运输缺陷有关。患者-父母三人组的全外显子组测序 (WES) 已被证明是识别导致散发性 ALS (sALS) 的罕见从头遗传变异的有效策略。使用三重 WFS 方法，我们在一名早发 sALS 患者中发现了新的 RAPGEF2 变异（c.4069G>A，p.E1357K）。为了评估这种变异的致病作用，我们使用了患者来源的皮肤成纤维细胞和果蝇中 RAPGEF2-E1357K 突变蛋白的运动神经元特异性过表达。患者成纤维细胞表现出微管稳定性降低和微管网络形态缺陷。患者细胞中线粒体的细胞内分布、超微结构和功能也受到损害。果蝇运动神经元中 RAPGEF2 突变体的过度表达会降低轴突微管的稳定性，并破坏线粒体到远端轴突和神经肌肉接头 (NMI) 突触的分布。我们还发现，与对照细胞相比，患者成纤维细胞中促凋亡蛋白 BCL2 相关 X (BAX) 向线粒体的募集显着增加。最后，通过组蛋白脱乙酰酶 6 (HDAC6) 的药理学抑制来增加微管稳定性，可挽救线粒体和 BAX 的细胞内分布缺陷。总体而言，我们的数据表明，本研究中发现的 RAPGEF2 变异可以通过微管失调驱动 ALS 相关的致病作用。
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that is frequently linked to microtubule abnormalities and mitochondrial trafficking defects. Whole exome sequencing (WES) of patient-parent trios has proven to be an efficient strategy for identifying rare de novo genetic variants responsible for sporadic ALS (sALS). Using a trio-WFS approach, we identified a de novo RAPGEF2 variant (c.4069G>A, p.E1357K) in a patient with early-onset sALS. To assess the pathogenic effects of this variant, we have used patient-derived skin fibroblasts and motor neuron-specific overexpression of the RAPGEF2-E1357K mutant protein in Drosophila. Patient fibroblasts display reduced microtubule stability and defective microtubule network morphology. The intracellular distribution, ultrastructure, and function of mitochondria are also impaired in patient cells. Overexpression of the RAPGEF2 mutant in Drosophila motor neurons reduces the stability of axonal microtubules and disrupts the distribution of mitochondria to distal axons and neuromuscular junction (NMI) synapses. We also show that the recruitment of the pro-apoptotic protein BCL2-associated X (BAX) to mitochondria is significantly increased in patient fibroblasts compared with control cells. Finally, increasing microtubule stability through pharmacological inhibition of histone deacetylase 6 (HDAC6) rescues defects in the intracellular distribution of mitochondria and BAX. Overall, our data suggest that the RAPGEF2 variant identified in this study can drive ALS-related pathogenic effects through microtubule dysregulation.