Miyazaki Yu, Du Xiaofei, Muramatsu Shin-Ichi, Gomez Christopher M
Department of Neurology, University of Chicago, Chicago, IL 60637, USA.
Division of Neurology, Department of Medicine, Jichi Medical University, Tochigi 3290498, Japan. Center for Gene and Cell Therapy, Institute of Medical Science, University of Tokyo, Tokyo 1088639, Japan.
Sci Transl Med. 2016 Jul 13;8(347):347ra94. doi: 10.1126/scitranslmed.aaf5660.
Spinocerebellar ataxia type 6 (SCA6) is a dominantly inherited neurodegenerative disease characterized by slowly progressive ataxia and Purkinje cell degeneration. SCA6 is caused by a polyglutamine repeat expansion within a second CACNA1A gene product, α1ACT. α1ACT expression is under the control of an internal ribosomal entry site (IRES) present within the CACNA1A coding region. Whereas SCA6 allele knock-in mice show indistinguishable phenotypes from wild-type littermates, expression of SCA6-associated α1ACT (α1ACTSCA6) driven by a Purkinje cell-specific promoter in mice produces slowly progressive ataxia and cerebellar atrophy. We developed an early-onset SCA6 mouse model using an adeno-associated virus (AAV)-based gene delivery system to ectopically express CACNA1A IRES-driven α1ACTSCA6 to test the potential of CACNA1A IRES-targeting therapies. Mice expressing AAV9-mediated CACNA1A IRES-driven α1ACTSCA6 exhibited early-onset ataxia, motor deficits, and Purkinje cell degeneration. We identified miR-3191-5p as a microRNA (miRNA) that targeted CACNA1A IRES and preferentially inhibited the CACNA1A IRES-driven translation of α1ACT in an Argonaute 4 (Ago4)-dependent manner. We found that eukaryotic initiation factors (eIFs), eIF4AII and eIF4GII, interacted with the CACNA1A IRES to enhance α1ACT translation. Ago4-bound miR-3191-5p blocked the interaction of eIF4AII and eIF4GII with the CACNA1A IRES, attenuating IRES-driven α1ACT translation. Furthermore, AAV9-mediated delivery of miR-3191-5p protected mice from the ataxia, motor deficits, and Purkinje cell degeneration caused by CACNA1A IRES-driven α1ACTSCA6 We have established proof of principle that viral delivery of an miRNA can rescue a disease phenotype through modulation of cellular IRES activity in a mouse model.
6型脊髓小脑共济失调（SCA6）是一种常染色体显性遗传的神经退行性疾病，其特征为缓慢进展的共济失调和浦肯野细胞变性。SCA6由第二个CACNA1A基因产物α1ACT内的多聚谷氨酰胺重复序列扩增引起。α1ACT的表达受CACNA1A编码区内存在的内部核糖体进入位点（IRES）控制。虽然SCA6等位基因敲入小鼠与野生型同窝小鼠表现出难以区分的表型，但在小鼠中由浦肯野细胞特异性启动子驱动的SCA6相关α1ACT（α1ACTSCA6）的表达会导致缓慢进展的共济失调和小脑萎缩。我们使用基于腺相关病毒（AAV）的基因递送系统开发了一种早发性SCA6小鼠模型，以异位表达CACNA1A IRES驱动的α1ACTSCA6，从而测试靶向CACNA1A IRES疗法的潜力。表达AAV9介导的CACNA1A IRES驱动的α1ACTSCA6的小鼠表现出早发性共济失调、运动缺陷和浦肯野细胞变性。我们鉴定出miR-3191-5p是一种靶向CACNA1A IRES的微小RNA（miRNA），并以AGO4依赖的方式优先抑制CACNA1A IRES驱动的α1ACT翻译。我们发现真核起始因子（eIFs），即eIF4AII和eIF4GII，与CACNA1A IRES相互作用以增强α1ACT翻译。与AGO4结合的miR-3191-5p阻断了eIF4AII和eIF4GII与CACNA1A IRES的相互作用，减弱了IRES驱动的α1ACT翻译。此外，AAV9介导的miR-3191-5p递送保护小鼠免受CACNA1A IRES驱动的α1ACTSCA6所导致的共济失调、运动缺陷和浦肯野细胞变性。我们已经建立了原理证明：在小鼠模型中，通过调节细胞IRES活性，病毒递送miRNA可以挽救疾病表型。
