Pathogenesis of Neurodegenerative Diseases Caused by Mutations in Senataxin

描述（由申请人提供）：本申请的目标是了解Sen1解旋酶在RNA聚合酶II （PolII）转录物终止中的作用，并确定Sen1人类同源基因Senataxin （SETX）的突变如何导致2型共济失调-动眼-失用症（AOA2）和4型肌萎缩性侧索硬化症（ALS4）的神经退行性疾病。基于酵母模型，我们提出SETX突变通过破坏转录终止导致人类疾病。因此，我们想要确定Sen1/SETX解旋酶在转录终止中的精确功能，以及这些功能在疾病中被破坏的方式。该基金的第一个目标是进一步了解酵母Sen1在转录终止中的作用以及转录终止机制的其他组成部分对其的调控。我们将确定Sen1是通过直接作用终止PolII，还是通过其对小核仁核糖核蛋白（snoRNP）组装的影响间接终止PolII。我们将通过分析在次优细胞Sen1水平下的snoRNP颗粒的蛋白质和RNA组成来实现这一目标。接下来，我们将确定Sen1突变导致细胞死亡的机制。这将通过减轻snoRNA信息读入下游必需基因的启动子阻断效应来实现。如果目前被接受的假设是正确的，这应该会抑制与Sen1缺失相关的致死率。随后，我们将探讨磷酸化在Sen1调控中的作用。第二个目标是确定酵母Sen1和人类SETX功能的共同特征。我们将亲和纯化SETX并鉴定与之相关的蛋白质和转录本。我们还将探讨这些关联是如何通过磷酸化调节的。第三个目标是确定人类SETX突变导致疾病的机制。我们将定义SETX突变如何改变其蛋白质和RNA结合特性，并检查SETX相关蛋白质的突变是否会导致与SETX突变相似的细胞代谢缺陷。我们相信上述目标的实现将有助于更好地理解转录终止的调控和神经退行性疾病的发病机制。获得的关于SETX功能的信息可能允许我们设计新的策略来逆转神经退行性疾病的进展，如ALS4和AOA2。我们的工作将研究Senataxin蛋白SETX的突变如何导致2型共济失调性动眼肌失用症（AOA2）和4型肌萎缩性侧索硬化症（ALS4）的神经退行性疾病。所获得的知识可能为逆转神经退行性疾病的进展提供新的策略。

DESCRIPTION (provided by applicant): The goal of this proposal is to understand the role of the Sen1 helicase in termination of RNA Polymerase II (PolII) transcripts and to establish how mutations in the human homolog of Sen1, Senataxin (SETX), cause the neurodegenerative disorders Ataxia-Oculomotor-Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis Type 4 (ALS4). Based on the yeast model, we propose that SETX mutations cause human disease through disruption of transcriptional termination. Thus, we want to define the precise function of the Sen1/SETX helicases in transcriptional termination and the manner in which these functions are disrupted in disease. The first aim of the grant is to learn more about the role of yeast Sen1 in transcription termination and its regulation by other components of the transcription termination machinery. We will determine whether Sen1 terminates PolII by direct action, or indirectly via its effect on small nucleolar ribonucleoprotein (snoRNP) assembly. We will accomplish this via analysis of the protein and RNA composition of the snoRNP particles under suboptimal cellular Sen1 levels. Next, we will identify the mechanism of the cell death caused by mutations of Sen1. This will be accomplished through alleviating the promoter occlusion effect of snoRNA messages reading into downstream essential genes. If the currently accepted hypothesis is correct, this should suppress the lethality associated with Sen1 loss. Subsequently we will explore the role of phosphorylation in the regulation of Sen1. The second aim is to identify the shared characteristics of yeast Sen1 and human SETX function. We will affinity purify SETX and identify the proteins and transcripts that associate with it. We will also explore how these associations are regulated by phosphorylation. The third aim is to identify the mechanism by which mutations in human SETX cause disease. We will define how mutations in SETX change its protein and RNA binding properties and examine whether mutations in proteins that SETX associates with cause similar defects in cellular metabolism as mutations in SETX. We believe that accomplishment of the goals outlined above will lead to a better understanding of the regulation of transcriptional termination and the pathogenesis of neurodegenerative disorders. The information obtained about the function of SETX might allow us to devise new strategies to reverse the progression of neurodegenerative diseases like ALS4 and AOA2. Project Narrative Our work will investigate how mutations in the Senataxin protein, SETX, cause the neurodegenerative disorders Ataxia Oculomotor Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis Type 4 (ALS4). The knowledge gained might provide new strategies to reverse the progression of neurodegenerative diseases.