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(TOR1A)蛋白中谷氨酸缺失的常染色体显性遗传引起，由于密码子或氨基酸缺失，通常被称为GAG或E突变。肌张力障碍背后的中枢神经系统异常知之甚少，功能成像和临床电生理学研究表明，整个运动回路的一系列结构都有异常。更具体的洞察应该来自对负责任的基因变化的理解。TorsinA是AAA+ATPase家族的成员，发现于内质网和核膜的管腔中。它的表达无处不在，已知的？E-突变酶不能将Torsin A从围产期致死的动物中拯救出来，这表明该突变缺乏Torsin A正常提供的基本活性。然而，TorsinA的具体功能差异很大，尽管自该蛋白首次被描述并与肌张力障碍有关以来，已有十多年的事实，但仍未得到很好的定义。这种缺乏洞察力正在成为开发有针对性和有效的肌张力障碍治疗方法的主要障碍。我们提出了一系列实验，旨在阐明TorsinA在培养的人类细胞中的细胞功能和与疾病相关的功能障碍。我们将建立在初步数据的基础上，表明TorsinA在内膜系统中的分布是受调节的，并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与内质网膜的结合基础及其在该细胞器中的分布和保持，(2)描述相互作用蛋白LULL1(TOR1IP2)控制TorsinA在内质网和核膜之间分布的机制，(3)利用细胞和生化分析来探索TorsinA在已知底物上的作用，以及(4)确定与疾病相关的突变如何影响TorsinA的结构和功能。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肌张力障碍的病因学提供了洞察力。 公共卫生相关性：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蛋白缺失的常染色体显性遗传引起。TorsinA的功能尚不清楚。我们的研究将确定这种重要酶的细胞作用和调节，从而为开发针对肌张力障碍的靶向疗法打开大门。肌张力障碍是继帕金森氏症和震颤之后的第三大常见运动障碍。

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. PUBLIC HEALTH RELEVANCE: Early-onset (DYT1) torsion dystonia is a devastating non-degenerative neurological movement disorder caused by autosomal dominant inheritance of a deletion in the protein torsinA. The function of torsinA is unknown. Our studies will define the cellular role and regulation of this important enzyme, thus opening the door to the development of targeted therapeutics for dystonia, which after Parkinson's disease and tremor is the is the third most common movement disorder in the US.