Defining the molecular drivers and modulators of MATR3 proteinopathy implicated in amyotrophic lateral sclerosis and frontotemporal dementia

项目摘要 Matrin-3（MATR 3）是一种普遍存在的RNA结合蛋白，其被预测为在很大程度上是无序的。突变 MATR 3与肌萎缩性侧索硬化症（ALS）和额颞叶痴呆（FTD）有关，而野生型MATR 3与肌萎缩性侧索硬化症（ALS）和额颞叶痴呆（FTD）有关。 在散发性ALS中观察到MATR 3型聚集体。然而，目前还不清楚致病性突变如何 干扰MATR 3毒性和溶解度。此外，RNA结合可以调节细胞的液-液相分离。 MATR 3.虽然MATR 3进入相分离的核仁，但不清楚MATR 3是核分裂的驱动者还是客户端。 相分离虽然MATR 3不像其他ALS RNA结合蛋白那样错误定位于细胞质，但它 可能会错误定位在细胞核内。这些发现被啮齿类动物的复杂性所混淆， 因此，开发MATR 3蛋白质病的简单模型以了解MATR 3蛋白质病的机制是至关重要的。 聚集的分子驱动器和改性剂。这一提议的驱动假设是， 与RNA结合减少相结合的突变是MATR 3聚集、错误定位和 毒性是ALS病理学的主要贡献者。一个多方面的方法酵母，纯蛋白 生物化学和哺乳动物神经元系统将用于研究MATR 3毒性的驱动因素。每个 这些系统的关键优势将共同揭示ALS/FTD的机制驱动因素和调节因素 发病机制初步数据表明，简单的酵母和纯蛋白质生化系统可以使用 阐明MATR 3毒性和聚集的分子驱动和修饰。核隔室 将使用SH-SY 5 Y细胞中的邻近标记和MATR 3的错误定位来鉴定MATR 3占据的细胞。 将被评估的扰动。最后，MATR 3的分子驱动因子是否可以引起ALS病理学 将在原代小鼠神经元中进行研究。具体而言，该提案将解决两个目标：1）定义 MATR 3毒性和聚集的分子决定因素和2）研究MATR 3病理学和定位 到神经元的核区。这些研究将为ALS/FTD的基础提供新的见解。 已经成立了一个指导小组，以确保在生物化学、液-液 相分离和神经科学，这将为学术研究生涯提供良好的准备。

Project Summary Matrin-3 (MATR3) is a ubiquitous RNA-binding protein which is predicted to be largely disordered. Mutations in MATR3 have been linked to amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) and wild- type MATR3 aggregates have been observed in sporadic ALS. However, it is unclear how pathogenic mutations perturb MATR3 toxicity and solubility. Additionally, RNA-binding may modulate liquid-liquid phase separation of MATR3. Although MATR3 enters the phase separated nucleolus, it is unclear if MATR3 is a driver or client of phase separation. While MATR3 does not mislocalize to the cytoplasm like other ALS RNA-binding proteins, it may mislocalize within the nucleus. These findings have been confounded by the complexities of rodent and primary neuron systems, so it is crucial to develop simple models of MATR3 proteinopathy to understand the molecular drivers and modifiers of aggregation. The driving hypothesis of this proposal is that pathogenic mutations in combination with diminished RNA binding are drivers of MATR3 aggregation, mislocalization, and toxicity which are major contributors to ALS pathology. A multifaceted approach of yeast, pure protein biochemistry, and mammalian neuronal systems will be used to investigate the drivers of MATR3 toxicity. Each of these systems has key benefits which together will uncover the mechanistic drivers and modifiers of ALS/FTD pathogenesis. Preliminary data suggests that simple yeast and pure protein biochemistry systems can be used to elucidate the molecular drivers and modifiers of MATR3 toxicity and aggregation. Nuclear compartments occupied by MATR3 will be identified using proximity labeling in SH-SY5Y cells, and mislocalization of MATR3 upon perturbation will be assessed. Finally, whether the molecular drivers of MATR3 can elicit ALS pathology will be investigated in primary mouse neurons. Specifically, this proposal will address two aims: 1) Define the molecular determinants of MATR3 toxicity and aggregation and 2) Investigate MATR3 pathology and localization to nuclear compartments in neurons. These studies will provide new insights into the underpinnings of ALS/FTD. A mentoring team has been established that will ensure key training opportunities in biochemistry, liquid-liquid phase separation, and neuroscience, which will provide excellent preparation for an academic research career.