Dissecting the Molecular Determinants of Cytoplasmic Gain of Function Pathomechanisms in TDP-43-linked Amyotrophic Lateral Sclerosis

剖析 TDP-43 相关肌萎缩侧索硬化症细胞质功能获得的分子决定因素

• 批准号: 2749409
• 金额: --
• 依托单位: King's College London
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2749409
• 关键词: Dissecting; Molecular; Determinants; Cytoplasmic; Gain

Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disease that leads to motor neuron loss. In vast majority of the cases (~95%), the RNA-binding protein TDP-43 is mislocalised and accumulates into cytoplasmic aggregates, a hallmark pathology of ALS (Neumann et al., 2006; Ling et al., 2013). As under physiological conditions TDP-43 is found in the nucleus and plays a significant role RNA metabolism, it's loss of function in the nucleus has been associated with significant alterations in RNA splicing and its liquid-liquid phase separation (LLPS) function (Polymenidou et al., 2011; Tollervey et al., 2011). On the other hand, evidence also supports cytoplasmic gain-of-function pathomechanisms for TDP-43 in disease (Suk and Rousseaux, 2020).For example, ALS-linked TDP-43 mutants can cause motor neuron degeneration in mice without aggregation or loss of nuclear TDP-43. Although studies have begun to unveil the molecular basis for TDP-43 RNA-binding and LLPS in cytoplasm, the pathomolecular determinants of cytoplasmic gain of functions in TDP-43-linked neurodegeneration have yet to be determined (Schmidt et al., 2019; Conicella et al., 2020; Hallegger et al., 2021). We propose generating three different novel mouse embryonic stem (mES) cell models to study the contribution of different disease associated properties of human cytoplasmic TDP-43, expressed in physiological levels.Aim of the investigation (up to 150 words)State primary research question and where appropriate the primary hypotheses being testedTDP-43 cytoplasmic aggregation is a key hallmark pathology of ALS. However, the contribution of different mechanisms by which mislocalised TDP-43 contributes to neurodegeneration is yet to be clearly elucidated. In this project we aim to dissect the roles of RNA-binding, LLPS and oligomerisation in TDP-43-linked ALS using novel TDP-43 mutant mouse embryonic stem (mES) cell models. In further studies, we aim to use these cells to develop in vivo mouse models to dissect neurodegeneration in a full animal context.Aim 1: Characterising novel CRISPR-edited mES cells to study TDP-43 linked ALSAim 2: Dissecting the cytoplasmic gain-of function TDP-43 pathomechanisms in novel CRISPR-edited mES cell derived neuronsAim 3: Determining cytoplasmic gain-of function TDP-43 pathomechanisms in novel and physiologically relevant in-vivo models for TDP-43 linked ALS

肌萎缩侧索硬化症(ALS)是一种成人起病的神经退行性疾病，导致运动神经元丢失。在绝大多数病例(~95%)中，RNA结合蛋白TDP-43被错误定位并积聚成细胞质聚集体，这是ALS的一种标志性病理(Neumann等人，2006年；Ling等人，2013年)。由于在生理条件下，TDP-43在细胞核中被发现并在RNA代谢中发挥重要作用，它在核中的功能丧失与RNA剪接及其液-液相分离(LLP)功能的显著变化有关(Polymenidou等人，2011年；Tollervey等人，2011年)。另一方面，证据也支持TDP-43在疾病中的细胞质功能增强病理机制(Suk和Rousseaux，2020)。例如，ALS连锁的TDP-43突变可以导致小鼠运动神经元退化，而不会聚集或丢失核TDP-43。尽管研究已经开始揭示TDP-43 RNA结合和细胞质中LLP的分子基础，但TDP-43连锁神经变性中细胞质功能获得的病理分子决定因素尚未确定(Schmidt等人，2019年；Conicella等人，2020年；Hregger等人，2021年)。我们建议建立三个不同的新的小鼠胚胎干细胞(MES)细胞模型，以研究人类细胞质TDP-43在生理水平表达的不同疾病相关特性的贡献。目的调查(长达150字)陈述主要研究问题，并在适当的情况下检验主要假设TDP-43细胞质聚集是ALS的关键标志病理学。然而，错位的TDP-43导致神经退行性变的不同机制的作用尚不清楚。在这个项目中，我们的目标是使用新的TDP-43突变的小鼠胚胎干细胞(MES)细胞模型来剖析RNA结合、LLP和寡聚在TDP-43连锁的ALS中的作用。在进一步的研究中，我们的目标是利用这些细胞在体内建立小鼠模型，在完整的动物环境下解剖神经变性。目标1：表征新的CRISPR编辑的MES细胞，以研究TDP-43连接的ALS。2：解剖新的CRISPR编辑的MES细胞来源的神经元的细胞质功能增益-TDP-43的病理机制。目标3：在TDP-43连接的ALS的新的和生理相关的体内模型中，确定TDP-43的细胞质功能增益-病理机制