The interaction between ATP13A2 and alpha-synuclein in mice

描述(申请人提供)：随着与帕金森氏症相关的新基因的不断发现，该领域的一个迫切问题是了解这些基因是否相互作用导致帕金森氏病的基本特征的发展。这种信息的缺乏是一个重要的问题，因为它限制了对帕金森病潜在共同机制的了解，从而也限制了新的疾病修改治疗方法的开发。长期目标是通过确定能够预防或显著减缓疾病进展的新的治疗靶点来改善帕金森氏症的治疗结果。这一特殊应用的目的是确定遗传基因座ATP13A2(PARK9)和α-突触核蛋白(PARK1)是如何相互作用的，特别是ATP13A2功能丧失如何影响小鼠的α-突触核蛋白毒性。ATP13A2缺失和α-突触核蛋白过表达的小鼠都表现出行为损伤和解剖病理方面的特征，这让人想起帕金森氏症，突出了它们的用处。中心假说是ATP13A2功能丧失增强了α-突触核蛋白的毒性，导致体内加速的多巴胺能细胞丢失和功能损伤。这一假设是在申请者实验室的初步研究和发表的研究的基础上提出的。这项拟议研究的基本原理是，一旦确定了与细胞和行为功能障碍相关的病理机制，就可以开始开发潜在的治疗方法。在初步数据的指导下，这一假说将通过追求两个具体目标来检验：1)确定ATP13A2缺失与体内1-突触核蛋白负荷增加之间的相互作用的功能效应。2)验证ATP13A2功能丧失联合α-突触核蛋白高表达导致溶酶体功能障碍、α-突触核蛋白异常蓄积增加、黑质纹状体多巴胺神经元加速死亡的假说。在第一个目标中，将申请人实验室培育的ATP13A2缺失小鼠与过度表达α-突触核蛋白的小鼠杂交，以产生过度表达α-突触核蛋白的双突变ATP13A2缺失小鼠。这些小鼠将在申请者实验室设计和建立的测试中，随着年龄的增长接受行为障碍的测试。在第二个目标中，将结合免疫组织化学技术、神经化学分析和体视学技术来测量多个年龄段的脑溶酶体蛋白、α-突触核蛋白和黑质纹状体病理。提出的方法是创新的，因为它专注于体内环境中与帕金森病相关的两个遗传位点之间的相互作用。这项拟议的研究具有重要意义，因为它是一系列研究的第一步，预计将导致识别导致帕金森病发生的机制，并提供一个模型，以测试针对这些机制的潜在疗法。

DESCRIPTION (provided by applicant): With the continued identification of novel genes associated with Parkinsonism an imperative question in the field is to know whether these genes interact to lead to the development of the cardinal features of Parkinson's disease. This lack of information is an important problem because it limits the understanding of potential common mechanisms underlying Parkinson's disease thus also limiting the development of novel disease-modifying treatments. The long-term goal is to improve treatment outcomes in Parkinson's disease by identifying novel therapeutic targets that can prevent or significantly slow the progression of the disease. The objective in this particular application is to determine how the genetic loci ATP13A2 (PARK9) and alpha-synuclein (PARK1) interact, specifically how does the loss of function of ATP13A2 affect alpha-synuclein toxicity in mice. ATP13A2 null and alpha-synuclein overexpressing mice each show aspects of behavioral impairment and anatomical pathology reminiscent of that seen in Parkinson's disease highlighting their usefulness. The central hypothesis is that loss of function of ATP13A2 enhances alpha-synuclein toxicity leading to accelerated dopaminergic cell loss and functional impairments in vivo. The hypothesis has been formulated on the basis of preliminary and published studies produced by the applicants' laboratories. The rationale for the proposed research is that once the pathologic mechanisms associated with cellular and behavioral dysfunction are identified, the development of potential therapies can be initiated. Guided by preliminary data this hypothesis will be tested by pursuing two specific aims: 1) Determine the functional effects of the interaction between loss of ATP13A2 and enhanced 1-synuclein protein burden in vivo. 2) To test the hypothesis that loss of function of ATP13A2 combined with overexpression of alpha-synuclein in mice leads to lysosomal dysfunction, increased abnormal accumulation of alpha-synuclein, and an acceleration of nigrostriatal dopamine neuron death in vivo. In the first aim, ATP13A2 null mice generated by the applicants' laboratory and mice that overexpress alpha-synuclein will be crossbred to generate double mutant ATP13A2 null mice that overexpress alpha-synuclein. These mice will be tested for behavioral impairments as they age on tests designed and established in the applicants' laboratories. In aim two, a combination of immunohistochemical techniques, neurochemical analysis, and stereological techniques will be used to measure brain lysosomal proteins, alpha-synuclein, and nigrostriatal pathology at multiple ages. The proposed approach is innovative, because it focuses on the interaction between two genetic loci linked to Parkinsonism in an in vivo environment. The proposed research is significant because it is the first step in a line of research expected to lead to the identification of mechanisms contributing to the development of PD and provide a model to test potential therapeutics targeting these mechanisms.