The Role of Stress Granules in the Pathophysiology of TDP-43

描述（由申请人提供）：TDP-43最近被确定为散发性肌萎缩性侧索硬化症（ALS）和泛素包涵体额颞叶痴呆（FTLD- U）包涵体中积累的主要蛋白质之一。TDP-43生物学异常足以引起神经退行性疾病，因为TDP-43突变与家族性ALS有关，这表明TDP-43与疾病过程直接相关。这项拨款提案的工作将建立在我们发现TDP-43调节对压力的转化反应的基础上。神经元对压力作出反应，例如在神经退行性疾病中，通过进入转译停止状态。翻译沉默的mRNA被隔离在称为应激颗粒（SG）的内含物中，该内含物由mRNA和RNA结合蛋白（如TIA-1）组成。由SGs组成的内含物因其动态特性而引人注目。使SGs成核的蛋白质具有可逆聚集的能力，部分原因是它们含有易聚集的聚谷氨酰胺和朊病毒结构域。TDP-43也是一种RNA结合蛋白。我们的研究表明，TDP-43与SGs共定位，对于SG的形成至关重要。TDP-43的疾病相关突变似乎会促进SG的形成，相反，抑制SG形成的药物会迅速分解TDP-43内含物。我们的数据还表明，翻译抑制剂可以抑制甚至快速分散TDP-43内含物。我们假设SGs有助于TDP-43包裹体的形成，例如发生在FTLD-U和ALS中，并且抑制SGs可以抑制TDP-43包裹体的形成。这个提议的目标1将把我们的研究从细胞系转移到初级神经元。我们的初步工作已在细胞系如人BE-M17神经母细胞瘤细胞中进行，但SGs在神经元中的调节不同。我们将确定在初级皮层和脊髓运动神经元培养中刺激TDP-43包涵体形成的条件类型。我们还将确定TDP-43的突变如何影响包涵体的形成。目的2将确定TDP-43包裹体形成的结构域和TDP-43包裹体的抑制/分散。这项工作将确定TDP-43的条件和区域，人们可以针对这些条件和区域开发可以抑制或逆转TDP-43内含物的治疗方法，并可能治疗ALS和FTLD-U。

DESCRIPTION (provided by applicant): TDP-43 was recently identified as one of the major proteins that accumulate in inclusions in sporadic Amyotrophic Lateral Sclerosis (ALS) and in Fronto-temporal dementia with ubiquitin inclusions (FTLD- U). Abnormalities in TDP-43 biology are sufficient to cause neurodegenerative disease because mutations in TDP-43 are linked to familial ALS, which suggests that TDP-43 is linked directly to the disease process. The work in this grant proposal will build on our discovery that TDP-43 regulates the translational response to stress. Neurons respond to stresses, such as occur in neurodegenerative diseases, by entering a state of translational arrest. Translationally silenced mRNA is sequestered into inclusions termed stress granules (SG), which are composed of an mRNA and RNA binding proteins, such as TIA-1. Inclusions composed of SGs are intriguing because of their dynamic nature. The proteins that nucleate SGs have the ability to reversibly aggregate partly because they contain aggregation prone polyglutamine and prion domains. TDP-43 is also a RNA binding protein. Our research demonstrates that TDP-43 co-localizes with SGs, and is essential for SG formation. Disease related mutations in TDP-43 appear to enhance SG formation, and conversely, agents that inhibit SG formation rapidly disaggregate TDP-43 inclusions. Our data also suggests that translational inhibitors can suppress or even rapidly disperse TDP-43 inclusions. We hypothesize that SGs contribute to formation of TDP-43 inclusions, such as occur in FTLD-U and ALS, and that inhibiting SGs can inhibit formation of TDP-43 inclusions. Aim 1 of this proposal will move our work from cell lines to primary neurons. Our preliminary work has been performed in cell lines such as human BE-M17 neuroblastoma cells, but the regulation of SGs differs in neurons. We will determine the types of conditions that stimulate formation of TDP-43 inclusions in primary cortical and spinal motor neuron cultures. We will also determine how mutations in TDP-43 affect inclusion formation. Aim 2 will identify the structural domains of TDP-43 inclusion formation and suppression/dispersion of TDP-43 inclusions. This work will identify the conditions and domains on TDP-43 that one can target to develop therapeutic approaches that can suppress or reverse TDP-43 inclusions and possibly treat ALS and FTLD-U. PUBLIC HEALTH RELEVANCE: This project investigates TDP-43, which is one of the major proteins that accumulate in Amyotrophic Lateral Sclerosis (ALS) and in Fronto-temporal dementia with ubiquitin inclusions (FTLD-U). We will determine the types of conditions that stimulate aggregation of TDP-43 and novel methods to prevent or reverse TDP-43 aggregation.