Deranged calcium signaling and polyglutamine expansion disorders

 描述（由申请人提供）：该项目的广泛，长期目标是了解亨廷顿病（HD）神经变性的原因。在HD中，多聚谷氨酰胺扩增的亨廷顿蛋白（Htexp）通过知之甚少的机制导致早期突触功能障碍和最终的神经变性。我们先前发现，Httexp结合到肌醇1，4，5-三磷酸受体1（InsP 3R 1），并使其对InsP 3敏感。我们还证明了一种新的神经元钙库操纵的钙内流（nSOC）抑制剂改善了转基因HD果蝇的运动症状，并抑制了YAC 128 HD小鼠中棘神经元（MSN）的神经变性，这是HD中受影响的主要细胞类型。基于这些结果，我们提出InsP 3R 1的过度激活导致内质网（ER）钙水平持续降低，nSOC升高以及HD患者MSN中神经元和树突棘丢失。为了验证这一假设，我们将集中在以下目标：1。为了测试InsP 3R 1的作用，我们将使用反义寡核苷酸（ASO）敲低YAC 128 MSN中的InsP 3R 1表达。我们还将通过过表达代谢InsP 3的5 PP-RA磷酸酶来降低InsP 3的水平。为了更直接地测试nSOC的作用，我们将使用遗传策略敲除或敲低突触nSOC的调节因子STIM 2。将在YAC 128 HD小鼠模型的体外和体内实验中评价InsP 3R 1和STIM 2介导的信号传导的抑制作用。2.目的探讨σ 1受体（sigma 1 receptor，S1 R）在HD中的作用，并探讨S1 R作为HD治疗靶点的可能性。σ 1受体（S1 R）是一种ER驻留蛋白，其响应于ER应激和ER钙耗竭而被激活。激活的S1 R被认为可以促进神经保护和稳定InsP 3R功能，这可能会保护MSN及其突触连接。我们的初步数据表明，在老年YAC 128小鼠和人类HD患者的纹状体S1 R的上调。我们将研究S1 R介导的信号在HD折磨纹状体的变化，并调查HD MSN棘S1 R和ER Ca 2+信号之间的潜在关系。我们还将使用各种药理学和分子工具来验证S1 R作为HD的新治疗靶点。在这些研究中，我们将评估失调的ER Ca 2+信号传导、nSOC和S1 R在HD MSNs中突触丢失中的潜在作用。我们还将评估InsP 3R 1，STIM 2和S1 R作为HD的潜在治疗靶点。

 DESCRIPTION (provided by applicant): The broad, long-term objective of the project is to understand the causes of neurodegeneration in Huntington's disease (HD). In HD polyglutamine-expanded Huntingtin (Httexp) causes early synaptic dysfunction and eventual neurodegeneration through poorly understood mechanisms. We previously discovered that Httexp binds to and sensitizes inositol 1,4,5-triphosphate receptor 1 (InsP3R1) to InsP3. We also demonstrated that a novel inhibitor of neuronal store-operated calcium entry (nSOC) improved motor symptoms in transgenic HD flies and inhibited and neurodegeneration in YAC128 HD mouse medium spiny neurons (MSNs), the primary cell type affected in HD. Based on these results, we propose that excessive activation of InsP3R1 causes a persistent reduction in endoplasmic reticulum (ER) calcium levels, elevated nSOC and neuronal and dendritic spine loss in HD-afflicted MSNs. To test this hypothesis, we will focus on the following aims: 1. To evaluate the causes and importance of supranormal nSOC in synaptic loss in HD MSNs To test the role of InsP3R1, we will use antisense oligonucleotides (ASOs) to knockdown InsP3R1 expression in YAC128 MSNs. We will also reduce levels of InsP3 by overexpressing a 5PP-RA phosphatase that metabolizes InsP3. To test the role of nSOC more directly we will use genetic strategies to knockout or knockdown STIM2, a regulator of synaptic nSOC. The effects of suppression of InsP3R1- and STIM2- mediated signaling will be evaluated in vitro and in vivo in experiments with the YAC128 HD mouse model. 2. To investigate the role of sigma 1 receptor (S1R) in HD and to evaluate S1R as a potential therapeutic target. Sigma 1 receptor (S1R) is an ER-resident protein that is activated in response to ER stress and ER calcium depletion. Activated S1R is thought to promote neuroprotection and stabilize InsP3R function, which may protect MSNs and their synaptic connections. Our preliminary data indicates upregulation of S1R in the striatum of both aged YAC128 mice and human HD patients. We will study changes in S1R- mediated signaling in the HD-afflicted striatum and investigate the potential relationship between S1R and ER Ca2+ signaling in HD MSN spines. We will also use a variety of pharmacological and molecular tools to validate S1R as a novel therapeutic target for HD. In these studies we will evaluate potential roles of dysregulated ER Ca2+ signaling, nSOC and S1R in synaptic loss in HD MSNs. We will also appraise InsP3R1, STIM2 and S1R as potential therapeutic targets for HD.