Spinocerebellar ataxia type 12 iPSCs and PP2A dysregulation

 描述(申请人提供)：脊髓小脑性共济失调12型(SCA12)是一种进行性常染色体显性遗传的神经退行性疾病，由染色体5q32上CAG/CTG三核苷酸重复序列的扩大引起；疾病表型和致病突变最初由我们小组描述(Holmes等人，1999)。虽然这种疾病是印度最常见的SCA形式之一，世界各地都发现了分散的SCA12家系，但SCA12最有趣的方面可能是重复发生在PPP2R2B的假定启动子上，PPP2R2B是一种编码三聚体酶磷酸酶2A(PP2A)的调节亚基的基因。功能性PP2A由一个结构单元、两个催化单元之一和一个~30个调节亚基组成，调节亚基的N端区域用于将全酶靶向于特定的细胞内位点。PP2A的失调与阿尔茨海默病中tau的过度磷酸化以及其他多种神经退行性疾病直接相关。根据细胞过度表达模型的初步数据，我们推测，SCA12重复序列的扩张导致PPP2R2B亚型B？1表达增加，这种过度表达导致PP2A活性的失调和神经毒性。然而，目前还无法证实这些观察结果，因为人类SCA12脑材料不可用，PPP2R2B在白细胞或淋巴母细胞中也不表达。为了验证我们的假设，我们将使用SCA12患者皮肤活检组织中的成纤维细胞来产生诱导多能干细胞(IPSCs)(目标1)。然后，我们将确定突变对成纤维细胞和分化为前脑神经元的IPSCs中PPP2R2B表达和其他细胞特性的影响(目标2)。该项目对公众健康的潜在益处有三个方面：1)更好地了解重复DNA如何影响基因表达；2)更好地了解SCA12的发病机制，有可能发现治疗药物的靶点；3)对PP2A在神经退行性疾病发病机制中的作用有新的见解。

 DESCRIPTION (provided by applicant): Spinocerebellar ataxia type 12 (SCA12) is a progressive, autosomal dominant, neurodegenerative disorder caused by an expansion of a CAG/CTG trinucleotide repeat on chromosome 5q32; both the disease phenotype and the causative mutation were initially described by our group (Holmes et al, 1999). While the disease is one of most common forms of SCA in India, and scattered SCA12 pedigrees have been detected around the world, perhaps the most intriguing aspect of SCA12 is that the repeat falls in a putative promoter of PPP2R2B, a gene encoding ß regulatory subunits of the trimeric enzyme phosphatase 2A (PP2A). Functional PP2A consists of a structural unit, one of two catalytic units, and one of ~30 regulatory subunits, with the N-terminal region of the regulatory subunits serving to target the holoenzyme to specific intracellular sites. Dysregulation of PP2A has been directly linked to tau hyperphosphorylation in Alzheimer's disease, and to multiple other neurodegenerative diseases. We hypothesize, based on preliminary data from cell overexpression models, that the SCA12 repeat expansion leads to increased expression of PPP2R2B isoform Bß1, and that this overexpression leads to dysregulation of PP2A activity and neurotoxicity. However, it has not been possible to confirm these observations, as human SCA12 brain material is not available and PPP2R2B Is not expressed in leukocytes or lymphoblasts. To test our hypothesis, we will use fibroblasts from skin biopsies of patients with SCA12 to generate induced pluripotent stem cells (iPSCs)(Aim 1). We will then determine the effect of the mutation on PPP2R2B expression and other cellular properties in the fibroblasts and in the IPSCs differentiated into forebrain neurons (Aim 2). The potential public health benefits of this project are three fold: 1) a better understanding of how repetitive DNA can influence gene expression, 2) a better understanding of SCA12 pathogenesis, with the potential of detecting targets for therapeutic agents, and 3) new insight into the role of PP2A in the pathogenesis of neurodegenerative disease.