FUNCTIONAL ANALYSIS OF TORSIN A

描述（由申请人提供）：早发性（DYT1）扭转肌张力障碍是一种破坏性的非退行性神经运动障碍，由torsinA蛋白（TOR1A）中谷氨酸缺失的常染色体显性遗传引起，通常被称为？呕吐还是？E突变是因为密码子或氨基酸缺失。肌张力障碍背后的中枢神经系统异常尚不清楚，功能成像和临床电生理学研究表明，运动回路中的一系列结构都存在异常。更具体的见解应该来自对负责任的基因变化的理解。TorsinA是在内质网管腔和核膜中发现的AAA+ atp酶家族的一员。它的表达无处不在，而已知的失败？将torsinA敲除的动物从围产期死亡中拯救出来的e -突变酶表明，这种突变体缺乏torsinA通常提供的任何必需活性。然而，尽管自该蛋白首次被描述并与肌张力障碍联系在一起已有十多年，但归因于torsinA的特定功能差异很大，并且尚未得到很好的定义。对肌张力障碍缺乏认识是开发有针对性和有效治疗方法的主要障碍。我们提出了一组旨在阐明培养的人细胞中torsinA的细胞功能和疾病相关功能障碍的实验。我们将建立在初步数据的基础上，这些数据表明，torsinA在膜系统内的分布是受调节的，并且可能在确定酶的活性方面发挥重要作用。该项目的具体目标是：(1)确定torsinA与内质网膜的关联基础及其在该细胞器中的分布和保留，(2)描述相互作用蛋白LULL1 （TOR1IP2）控制内质网和核膜之间torsinA分布的机制，(3)利用细胞和生化分析探索torsinA对已知底物的影响。(4)确定疾病相关突变如何影响torsinA结构和功能。这些研究具有广泛的相关性，因为它们解决了调节细胞内蛋白质定位的潜在新方法，同时也为DYT1肌张力障碍的病因提供了见解。

DESCRIPTION (provided by applicant): Early-onset (DYT1) torsion dystonia is a devastating non-degenerative neurological movement disorder caused by autosomal dominant inheritance of a glutamic acid deletion in the protein torsinA (TOR1A), frequently referred to as the ?GAG or ?E mutation because of the deleted codon or amino acid. The CNS abnormalities underlying dystonia are poorly understood, with functional imaging and clinical electrophysiology studies suggesting abnormalities in a range of structures throughout the motor circuit. More specific insight should come from understanding the responsible genetic change. TorsinA is a member of the AAA+ family of ATPases found in the lumen of the endoplasmic reticulum and nuclear envelope. It is expressed ubiquitously, and the known failure of ?E-mutant enzyme to rescue torsinA knock-out animals from perinatal lethality suggests that this mutant lacks whatever essential activity torsinA normally provides. However, the specific functions ascribed to torsinA vary widely and are not well defined despite the fact that it has been more than a decade since the protein was first described and linked to dystonia. This lack of insight is creating a major roadblock in efforts to develop targeted and effective treatments for dystonia. We propose a set of experiments aimed at clarifying the cellular function and disease-linked dysfunction of torsinA in cultured human cells. We will build on preliminary data showing that the distribution of torsinA within the endomembrane system is regulated and likely to play an important role in defining the enzyme's activity. The specific aims of the project are (1) to define the basis for association of torsinA with the endoplasmic reticulum membrane and its distribution and retention in this organelle, (2) to delineate the mechanism by which an interacting protein LULL1 (TOR1IP2) controls the distribution of torsinA between the endoplasmic reticulum and nuclear envelope, (3) to explore the effects of torsinA on known substrates using cellular and biochemical assays, and (4) to determine how disease-associated mutations affect torsinA structure and function. These studies are broadly relevant because they address potentially novel means of regulating the localization of proteins within cells, while also providing insight into the etiology of DYT1 dystonia.