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

项目摘要 Matrin-3(MATR3)是一种普遍存在的RNA结合蛋白，被预测在很大程度上是无序的。基因突变 MATR3与肌萎缩侧索硬化症(ALS)、额颞叶痴呆(FTD)和野生型 在散发性ALS中观察到MATR3型聚集体。然而，目前尚不清楚致病突变是如何 扰动MATR3的毒性和溶解性。此外，RNA结合可能调节液液相分离。 材料3.虽然MATR3进入了相分离的核仁，但目前还不清楚MATR3是驱动还是客户 相分离。虽然MATR3不像其他ALS RNA结合蛋白那样错误定位到细胞质，但它 可能在细胞核内错位。这些发现被啮齿动物和 因此，建立简单的MATR3蛋白病变模型以了解 聚集的分子驱动因素和修饰者。这一提议的驱动假设是致病的 突变和RNA结合减弱是MATR3聚集、错误定位和 毒性是ALS病理的主要贡献者。酵母、纯蛋白的多方面提取方法 生物化学和哺乳动物神经系统将被用来研究MATR3毒性的驱动因素。每个人 这些系统的关键优势将共同揭示ALS/FTD的机械驱动因素和修饰物 发病机制。初步数据表明，可以使用简单的酵母和纯蛋白质生物化学系统。 阐明MATR3毒性和聚集的分子驱动因素和修饰物。核舱室 将通过SH-SY5Y细胞中的邻近标记和MATR3的错误定位来鉴定MATR3所占据的位置 将对扰动进行评估。最后，MATR3的分子驱动因素是否能引发ALS病理 将在原代小鼠神经元中进行研究。具体地说，这项提案将涉及两个目标：1)界定 MATR3毒性和聚集的分子决定因素以及2)MATR3的病理和定位研究 到神经元中的核隔间。这些研究将为ALS/FTD的基础提供新的见解。 已经建立了一个指导小组，以确保在生物化学、液体-液体方面的关键培训机会 相分离和神经科学，这将为学术研究生涯提供极好的准备。

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.