RNA Dysfunction in Selectively Vulnerable Populations in SCA7 Mice

项目摘要/摘要： 脊髓小脑性共济失调 7 型（SCA7）是一种常染色体显性遗传性神经退行性疾病，由以下原因引起： 继承了 ataxin-7 基因编码区的 CAG 重复扩增，从而产生聚谷氨酰胺 (polyQ) 扩展突变蛋白。 SCA7 与其他神经退行性疾病具有共同特征 由 PolyQ 扩展突变（例如亨廷顿病）引起。其中一个特征是退化 发生在选择性脆弱的神经群体中。在 SCA7 中，退化的群体包括浦肯野 细胞 (PC)、伯格曼胶质细胞 (BG) 和下橄榄神经元 (IO)，它们将攀爬纤维轴突发送到 PC 树突上的突触。使用 SCA7 动物模型，我们发现疾病基因表达 特别是在 BG、PC 和 IO 神经元中，影响 SCA7 疾病表型。有趣的是，使用 floxed- SCA7的polyQ ataxin-7小鼠模型我们观察到Cre重组酶在所有三种细胞类型中表达 大大延迟了症状的出现。综上所述，这些发现支持突变体的假设 这三种细胞类型中 ataxin-7 的表达会导致细胞相互作用功能失调，并且是一个关键因素 SCA7 小脑变性的介质。不幸的是，SCA7 的分子机制 这 3 种特定细胞类型的病理学尚未确定。我们还观察到 症状出现后抑制突变基因表达可以阻止疾病进展，但不能逆转 PC 或 BG 中的病理学。然而，在 SCA7 中，小脑的 IO 输入均减少并重新分配 老鼠。 IO-PC 突触病理学是通过抑制突变体来防止的唯一可检测到的异常 症状出现后的基因表达，表明 IO-PC 突触的丧失导致 SCA7 疾病 进展。由于 SCA7 小脑病理学涉及特定细胞类型之间相互作用的改变， 存在于中枢神经系统的不同区域，研究基因表达或蛋白质改变的标准方法 内容无法区分病理学中涉及的细胞类型特异性变化 针对选择性脆弱细胞的退化而发生的过程和反应性变化。至 为了解决这个问题，我们开发了从三种细胞中特异性分离 RNA 的技术 已知影响小脑病理学和运动行为的人群。在这个项目中，我们将使用这些 解决编码和非编码 RNA 中细胞类型特异性变化导致的假设的方法 在 SCA7 小鼠小脑中观察到的细胞功能障碍的特定模式。我们将进一步确定 通过诱导型 Cre- 抑制突变 ataxin-7 表达后，RNA 变化是可逆的 重组酶。这些发现驱动的实验具有高风险，但有可能产生关键成果 有关导致 PolyQ 疾病选择性脆弱性的分子机制的信息。

Project Summary/Abstract: Spinocerebellar ataxia type 7 (SCA7) is an autosomal dominant neurodegenerative disorder caused by inheriting a CAG repeat expansion in the coding region of the ataxin-7 gene, resulting in a polyglutamine (polyQ) expanded mutant protein. SCA7 has features in common with other neurodegenerative disorders caused by polyQ expansion mutations such as Huntington's Disease. One such feature is that degeneration occurs in selectively vulnerable neural populations. In SCA7 the populations that degenerate include Purkinje cells (PCs), Bergmann Glia (BG) and neurons of the inferior olive (IO) which send climbing fiber axons to synapse on PC dendrites. Using animal models of SCA7, we have shown that disease gene expression specifically in BG, PCs and IO neurons influence the SCA7 disease phenotype. Interestingly, using a floxed- polyQ ataxin-7 mouse model of SCA7 we observed that expression of Cre recombinase in all three cell types dramatically delayed symptom onset. Taken together, these findings support the hypothesis that mutant ataxin-7 expression in these three cell types contributes to dysfunctional cellular interactions and is a critical mediator of SCA7 cerebellar degeneration. Unfortunately, the molecular mechanisms responsible for SCA7 pathology in these 3 specific cell types have not yet been determined. We have also observed that suppression of mutant gene expression after symptom onset halts disease progression, but does not reverse pathology in PCs or BG. However, IO inputs to the cerebellum are both decreased and redistributed in SCA7 mice. IO-PC synapse pathology was the only detectable abnormality prevented by suppression of mutant gene expression after symptom onset, suggesting that loss of IO-PC synapses contributes to SCA7 disease progression. Since SCA7 cerebellar pathology involves altered interactions between specific cell types that reside in distinct regions of the CNS, standard approaches to the study of altered gene expression or protein content cannot distinguish between cell type specific changes mechanistically involved in the pathologic process and reactive changes that develop in response to the degeneration of selectively vulnerable cells. To address this issue, we have developed techniques to isolate RNA specifically from the three cellular populations known to impact cerebellar pathology and motor behavior. In this project, we will employ these methods to address the hypothesis that cell type specific changes in both coding and non-coding RNAs lead to the specific pattern of cellular dysfunction observed in the cerebellum of SCA7 mice. We will further identify which RNA changes are reversible following suppression of mutant ataxin-7 expression by inducible Cre- recombinase. These discovery driven experiments are high-risk, but have the potential to yield critical information regarding the molecular mechanisms responsible for selective vulnerability in a polyQ disorder.