Spinocerebellar Ataxia Type 2 Gene and Gene Product

脊髓小脑共济失调 2 型基因和基因产物

• 批准号: 8739997
• 负责人: Stefan M. PULST
• 金额: $ 47.34万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-02-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8739997
• 关键词: 22q; Address; Affect; Age; Aging; Agonist; Alleles; Alternative Splicing; Amyotrophic Lateral Sclerosis; Animals; Anus; Ataxia; Behavioral; Biochemical; Brain; Brain Stem; CAG repeat; Calcium; Calcium Channel; Cause of Death; Cell Death; Cell physiology; Cells; Cerebellar Nuclei; Cerebellum; Code; Collaborations; Dancing; Detection; Development; Disease; Disease Progression; Economic Burden; Equilibrium; Event; Fiber; Frequencies; Funding; Gene Expression Profile; Gene Mutation; Genes; Genetic; Grant; Health; Homeostasis; Human; ITPR1 gene; In Vitro; Inherited; Inositol; Knock-out; Knockout Mice; Lead; Malignant Neoplasms; Messenger RNA; Metabotropic Glutamate Receptors; Modeling; Molecular Profiling; Motor; Movement; Mus; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Neuroprotective Agents; Parkinson Disease; Parkinsonian Disorders; Pathogenesis; Pathway Analysis; Pathway interactions; Peat; Phenotype; Physiological; Population; Preparation; Prevalence; Process; Protein Isoforms; Proteins; Purkinje Cells; Research; Rodent; Rodent Model; Severities; Signal Transduction; Slice; Spinocerebellar Ataxias; Synapses; Techniques; Testing; Time; Transcript; Transgenic Model; Type 2 Spinocerebellar Ataxia; Western Blotting; base; behavior test; cell type; deep sequencing; human disease; in vivo; interdisciplinary approach; laser capture microdissection; motor neuron degeneration; mouse model; mutant; neurophysiology; novel; parkin gene/protein; polyglutamine; postnatal; promoter; public health relevance; receptor; research study; tripolyphosphate; voltage

DESCRIPTION (provided by applicant): Neurodegenerative diseases represent an ever-increasing societal and economic burden with WHO estimates indicating that they will replace cancer as the 2nd leading cause of death by 2040. Degenerative ataxias are a common form of neurodegeneration affecting cerebellar Purkinje cells (PCs) and other neurons in the CNS. Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant form of ataxia caused by expansion of a coding CAG repeat and belongs to the group of polyglutamine (polyQ) diseases. Mutations in the SCA2 (ATXN2) gene can also cause or contribute to the development of Parkinson disease and ALS, two common forms of neurodegeneration. Neither symptomatic nor neuroprotective agents have been identified for the treatment of human ataxias. In the previous funding period we developed a multidisciplinary approach to characterize several SCA2 mouse models combining morphologic, biochemical, physiologic, and behavioral techniques. We used the cerebellar slice preparation to show that a decrease in PC firing frequency closely mirrored the decline in motor function. Importantly, significant transcriptome and neurophysiological changes occurred before any dendritic or cell loss was apparent in the cerebellum. We discovered that mutant ATXN2 induced abnormal calcium release from the ER via enhanced interaction with the inositol-triphosphate receptor (ITPR1). Based on these findings, two specific aims are proposed. In Aim 1, we will test the hypothesis that the mGluR1-ITPR1 axis is hyperactive in cerebella from SCA2 mice expressing polyQ expanded ATXN2. We will use the cerebellar slice preparation to evaluate mGluR1 signaling and the relationship between PC firing and intracellular calcium. We will also test the mGluR1-ITPR1 axis in vivo by genetic interaction with mGluR1 haploinsufficient mouse lines. In aim 2, we will use transcriptome profiling at presymptomatic time points to identify novel genes involved in SCA2 pathogenesis. To increase sensitivity and the ability to identify changes in less abundantly PC-expressed genes we have established laser-capture microdissection (LCM) of cerebellar regions. Transcriptomes will be analyzed by GO and KEGG pathway analysis with a particular emphasis on mGluR1-ERK downstream targets. Changes in key genes will be verified by qPCR and ICC/ western blot analysis. We hypothesize that some expression changes will be homeostatic, but others will contribute to pathogenesis. We will differentiate between these alternatives by normalizing expression of the respective gene using AAV- transduction in vivo. Five genes will be chosen based on presence of early changes and presence of changes in multiple SCA2 mouse models. Mice will be evaluated by behavioral testing as well as analysis in the cerebellar slice. The proposed experiments address two significant questions relating to SCAs: the importance of intracellular calcium levels on PC function and survival and the identification of novel pathways that are shared across multiple SCA2 rodent models. Answers to these questions will help in the identification of new avenues towards treatments of SCA2 and other degenerative ataxias.

描述(申请人提供)：神经退行性疾病是一个不断增加的社会和经济负担，世卫组织估计，到2040年，它们将取代癌症成为第二大死亡原因。退行性共济失调是一种常见的神经变性形式，影响小脑浦肯野细胞(PC)和中枢神经系统的其他神经元。脊髓小脑型共济失调2(Sca2)是一种常染色体显性遗传性共济失调，由编码CAG重复序列的扩增引起，属于聚谷氨酰胺(PolyQ)病。SCA2(ATXN2)基因的突变也可能导致或导致帕金森病和ALS的发展，这两种常见的神经退化形式。目前还没有发现治疗人类共济失调的对症药物和神经保护剂。在之前的资助期间，我们开发了一种多学科的方法来表征几种SCA2小鼠模型，结合了形态、生化、生理学和行为技术。我们使用小脑切片准备来显示PC放电频率的降低与运动功能的下降密切相关。重要的是，显著的转录组和神经生理学变化发生在小脑中任何树突或细胞丢失之前。我们发现，突变型ATXN2通过增强与肌醇-三磷酸受体(ITPR1)的相互作用，诱导内质网异常钙释放。基于这些发现，本文提出了两个具体目标。在目标1中，我们将测试mGluR1-ITPR1轴在表达多Q扩展ATXN2的SCA2小鼠的小脑中过度活跃的假设。我们将使用小脑切片制备来评估mGluR1信号以及PC放电与细胞内钙的关系。我们还将通过与mGluR1单倍体缺陷小鼠株系的遗传相互作用，在体内测试mGluR1-ITPR1轴。在目标2中，我们将使用症状前时间点的转录组图谱来识别与SCA2发病相关的新基因。为了提高敏感性和识别PC表达较少的基因变化的能力，我们建立了小脑区域的激光捕获显微解剖(LCM)。转录本将通过GO和KEGG途径分析，特别强调mGluR1-ERK下游靶点。关键基因的改变将通过定量聚合酶链式反应和ICC/Western印迹分析来验证。我们假设有些基因表达的改变是动态平衡的，但另一些则有助于发病。我们将通过在体内使用AAV转导使各自基因的表达正常化来区分这些替代方案。根据早期变化的存在和在多个SCA2小鼠模型中存在的变化，将选择五个基因。小鼠将通过行为测试和小脑切片分析进行评估。拟议的实验解决了与SCA2相关的两个重要问题：细胞内钙水平对PC功能和生存的重要性，以及识别在多种SCA2啮齿动物模型中共享的新途径。对这些问题的回答将有助于确定治疗SCA2和其他退行性共济失调的新途径。