The impact of TDP-43 on translation and the response to axonal damage in amyotrophic lateral sclerosis

肌萎缩侧索硬化症(ALS)，又称运动神经元病(MND)，是一种破坏性的神经退行性疾病，导致MN进行性丧失，导致肌肉功能受损和瘫痪。肌萎缩侧索硬化症是无法治愈的，并会导致死亡，通常是由呼吸困难引起的，平均在确诊后3年内，一生中有大约1/400的风险。运动神经元(MN)是将信号从脊髓发送到肌肉的神经细胞。它们是体内最大的细胞之一，它们的细胞体位于脊髓内，其纤维(称为轴突)向外伸向肌肉。所有的细胞隔间，包括轴突，都需要新合成的蛋白质的持续供应才能正常运作。在某些情况下，例如，当对细胞损伤等不可预测的事件做出反应时，绝对需要新的不同的蛋白质来重建细胞功能和长期生存。这对轴突来说尤其具有挑战性，因为这些蛋白质的RNA是制造它们所必需的，可能需要在非常远的核中合成。多条证据表明，ALS疾病机制中的关键角色之一是一种名为TDP-43的蛋白质，它对于RNA特定运输到轴突中的不同位置以及细胞对应激和损伤的反应至关重要。在本篇论文中，我将测试TDP-43如何影响MNS对轴突损伤的反应，以及这一反应途径在ALS中的相关性。为此，我将把新的小鼠疾病模型和患者细胞系与最先进的生物学工具结合起来，以便研究这些非常特定的功能。特别是，我们已经开发出携带小鼠TDP-43基因特定突变的新小鼠品系，从而发展出ALS的关键特征。这些肌萎缩侧索硬化症的小鼠模型将使我们能够通过观察症状前期的小鼠和有明显肌萎缩侧索硬化症症状的小鼠，识别疾病进展过程中发生的分子和细胞变化。我们将使用一种创新和完全集成的方法，利用新的分子生物学和显微技术分析这些小鼠的疾病表型，以确定包括轴突在内的不同MN区域存在哪些类型的RNA和蛋白质，以及这些RNA和蛋白质如何对损伤做出反应。因此，我将能够通过以下研究来研究TDP-43在轴突损伤反应中引发的细胞变化：1.在轴突与细胞体分离的特殊培养皿中生长的MN；2.野生型和突变小鼠的轴突及其损伤反应；3.通过验证我们对来自患者诱导干细胞的MN的结果，并使用ALS患者捐赠的死后脑样本进行研究。综上所述，该项目将有助于了解TDP-43的变化如何影响MN的存活率。这一期待已久的信息对于开发有效的运动神经元疾病治疗方法至关重要。

Amyotrophic lateral sclerosis (ALS), also known as motor neuron disease (MND), is a devastating neurodegenerative disorder that causes progressive loss of MNs, leading to impaired muscle function and paralysis. ALS is incurable and leads to death, usually caused by the inability to breathe, on average only 3 years after diagnosis, with a lifetime risk of about 1 in 400. Motor neurons (MNs) are the nerve cells that send signals from the spinal cord to our muscles. They are amongst the largest cells in the body, with their cell soma located in the spinal cord and its fiber (called 'axon') projecting outside to the muscles. All cell compartments, including axons, need a constant supply of newly synthesised proteins in order to function properly. On some occasions, for example when responding to an unpredicted event as cell damage, new and different proteins are absolutely needed to re-establish cell functionality and long term survival. This is particularly challenging for axons, as the RNA for these proteins, which is essential to make them, may need to be synthesized very far away in the nucleus. Multiple lines of evidence indicate that one of the crucial players in the disease mechanism of ALS is a protein called TDP-43, which is important for the specific transport of RNA to different locations in the axons and in the response of cells to stress and damage.In this Fellowship, I will test how TDP-43 impacts on the response of MNs to damage in the axons, and the relevance of this response pathway in ALS. To do so, I will combine novel mouse models of disease and patient cell lines with state-of-the-art biological tools that will allow the investigation of these very specific functions.In particular, we have developed novel strains of mice that carry specific mutations in the mouse TDP-43 gene and, as a result, develop crucial features of ALS. These mouse models of ALS will allow us, by looking over time at pre-symptomatic mice and at mice with overt ALS symptoms, to identify the molecular and cellular changes occurring during disease progression. Using an innovative and fully integrated approach, we will analyse the disease phenotype of these mice with new molecular biology and microscopic techniques to identify which types of RNAs and proteins are present in different MN regions, including the axon, and how these change in response to damage. I will therefore be able to investigate the cellular alterations triggered by TDP-43 in axonal damage response by studying: 1. MNs grown in special culture dishes where the axons are separated from the cell bodies; 2. The axons and their damage response in wild type and mutant mice; and 3. By validating our results in MNs derived from patient-induced stem cells and using post mortem brain samples donated to research by ALS patients. In summary, this project will contribute to understand how changes in TDP-43 impacts on MN survival. This long-awaited information is essential to develop effective therapeutics for motor neuron disorders.