Evaluation of the role of RNA toxicity in SCA2 pathogenesis using genome editing in patient iPSCs

脊髓小脑性共济失调2型（SCA 2）是一种由CAG重复序列引起的破坏性神经退行性疾病 基因ataxin-2（ATXN 2）的扩增。CAG重复序列被翻译成多聚谷氨酰胺（polyQ）片段， ATXN 2突变蛋白具有神经毒性。目前的治疗努力集中在抑制 突变ATXN 2蛋白的表达，或靶向神经毒性的下游途径。我们和其他人 已经表明，含有CAG/CTG重复序列的突变体转录物也有助于几种疾病的发病机制， 重复疾病，包括亨廷顿病（HD）、亨廷顿病样2（HDL 2）和脊髓小脑 共济失调3型和8型（SCA 3和SCA 8）。因此，除了靶向突变蛋白的神经毒性外， 成功的治疗可能需要靶向突变转录物和/或突变转录物下游的途径 成绩单我们的初步证据支持这样的观点，即在SCA 2中，除了毒性ATXN 2蛋白， 有义ATXN 2 RNA和含有扩增的CUG重复序列的ATXN 2反义转录物（ATXN 2-AS）， 有助于SCA 2发病机制。因此，我们建议使用基因组编辑方法来修改正常和 将SCA 2 iPSC系转化为新的等基因iPSC系，其模拟SCA 2的蛋白质或RNA诱导机制 发病机制这些品系将用于（1）进一步检验SCA 2的新假设， ATXN 2/ATXN 2-AS转录物破坏SCA 2中的RNA加工，包括RNA输出、rRNA代谢和RNA合成。 剪接，和（2）确定RNA神经毒性对SCA 2发病机制的相对贡献。结果 从这项研究中获得的信息不仅有助于我们理解日益复杂的发病机制， SCA 2，但也有其他CAG/CTG重复疾病，其中双向编码的转录本和 蛋白质有助于病理学。这些结果将有助于指导SCA 2治疗的未来发展，以及 其他微卫星重复扩增疾病的治疗。

Spinocerebellar ataxia type 2 (SCA2) is a devastating neurodegenerative disease caused by a CAG repeat expansion in the gene ataxin-2 (ATXN2). The CAG repeat is translated into a polyglutamine (polyQ) tract in the mutant ATXN2 protein that has neurotoxic properties. Current therapeutic efforts are focused at suppressing the expression of the mutant ATXN2 protein, or targeting downstream pathways of neurotoxicity. We and others have shown that mutant CAG/CTG repeat-containing transcripts also contribute to pathogeneses of several repeat diseases, including Huntington's disease (HD), Huntington's disease-like 2 (HDL2) and spinocerebellar ataxia type 3 and 8 (SCA3 and SCA8). Therefore, in addition to targeting the neurotoxicity of mutant proteins, a successful therapy may require targeting the mutant transcripts and/or pathways downstream of the mutant transcripts. Our preliminary evidence supports the idea that in SCA2, in addition to toxic ATXN2 protein, both sense ATXN2 RNA and a transcript antisense to ATXN2 (ATXN2-AS) containing an expanded CUG repeat, contribute to SCA2 pathogenesis. We therefore propose to use genome editing approach to modify normal and SCA2 iPSC line into novel, isogenic iPSC lines that model either protein or RNA-induced mechanisms of SCA2 pathogenesis. These lines will be used to (1) further test the novel hypothesis of SCA2 by which mutant ATXN2/ATXN2-AS transcripts disrupt RNA processing in SCA2, including RNA export, rRNA metabolism and splicing, and (2) determine the relative contribution of RNA neurotoxicity to SCA2 pathogenesis. The results obtained from this study will not only facilitate our understanding of the increasingly complex pathogenesis of SCA2, but also that of other CAG/CTG repeat diseases in which both bi-directionally encoded transcripts and protein contribute to pathology. The results will help guide future development of SCA2 therapy, as well as therapies of other microsatellite repeat expansion diseases.