MOUSE MUSCLEBLIND MODEL FOR MYOTONIC DYSTROPHY

强直性肌营养不良小鼠肌盲模型

• 批准号: 6824697
• 负责人: MAURICE SCOTT SWANSON
• 金额: $ 29.83万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6824697
• 关键词: RNA splicing; adeno associated virus group; chloride channels; cooperative study; disease /disorder model; gene deletion mutation; gene expression; genetically modified animals; laboratory mouse; model design /development; molecular pathology; muscle proteins; myogenesis; myotonic dystrophy; phenotype; recombinant proteins; recombinant virus

The myotonic dystrophies (DM1 and DM2), which are the most common form of adult-onset muscular dystrophy, are autosomal dominant diseases with similar clinical presentations. Remarkably, DM1 and DM2 are caused by unstable microsatellite expansions in the untranslated regions of two different genes, DMPK and ZNF9. To explain how these non-coding expansion mutations lead to dominantly inherited neuromuscular disorders, we have proposed a toxic RNA model for the myotonic dystrophies. Transcription of the mutant DM1 (CTG)n and DM2 (CCTG)n alleles leads to the production of unusual RNA transcripts with (CUG)n and (CCUG)n repeat expansions. These expansions fold into stable double-stranded (ds) RNA structures that recruit and then sequester a family of dsRNA-binding factors, the muscleblind proteins. Because this toxic RNA model suggests that DM1 and DM2 diseases are due to loss of muscleblind protein function, we have derived muscleblind 1 (Mbnl1) knockout mice. This proposal is designed to test our working hypothesis that Mbnl1-/- knockout mice will be a useful model to examine underlying molecular mechanisms involved in myotonic dystrophy disease pathogenesis. First, we will characterize the Mbnl1-/-muscle phenotype and test the hypothesis that Mbnl1 is required for proper alternative splicing and function of the chloride channel CIC-1. Deficiency of this ion channel has been recently implicated as the cause of DM1- and DM2- associated myotonia. The stoichiometric relationship between toxic RNA and binding protein will be examined by breeding Mbnll knockout mice with lines of transgenic mice that express (CUG)n RNA at different levels. Second, the possibility that muscleblind proteins influence CIC-1 chloride channel levels by interacting with alternative splicing, and/or other, factors will be examined. Third, the hypothesis that the myotonia phenotype can berescued using recombinant adeno-associated virus mediated expression of wild type adult CIC-1 will be tested. Finally, we will investigate if additional disease-associated phenotypes result from deletion of the entire Mbnl1 gene, from tissue-specific Mbnl1 expression or from combinatorial loss of all three muscleblind (Mbnl1, Mbnl2/Mbnll, Mbnl3/Mblx) genes.

强直性肌营养不良症（DM1 和 DM2）是成人发病的肌营养不良症最常见的形式，是具有相似临床表现的常染色体显性遗传疾病。值得注意的是，DM1 和 DM2 是由两个不同基因 DMPK 和 ZNF9 的非翻译区不稳定的微卫星扩增引起的。为了解释这些非编码扩展突变如何导致显性遗传性神经肌肉疾病，我们提出了强直性肌营养不良的毒性 RNA 模型。突变体 DM1 (CTG)n 和 DM2 (CCTG)n 等位基因的转录导致产生具有 (CUG)n 和 (CCUG)n 重复扩展的异常 RNA 转录物。这些扩展折叠成稳定的双链 (ds) RNA 结构，招募并隔离一系列 dsRNA 结合因子，即肌肉盲蛋白。因为这种有毒的 RNA 模型表明 DM1 和 DM2 疾病是由于肌盲蛋白的丢失造成的 功能，我们衍生出了muscleblind 1（Mbnl1）基因敲除小鼠。该提案旨在测试我们的工作假设，即 Mbnl1-/- 敲除小鼠将成为检查强直性肌营养不良疾病发病机制的潜在分子机制的有用模型。首先，我们将表征 Mbnl1-/-肌肉表型，并测试 Mbnl1 是氯离子通道 CIC-1 的正确选择性剪接和功能所必需的假设。最近，该离子通道的缺陷被认为是 DM1 和 DM2 相关肌强直的原因。将通过用不同水平表达 (CUG)n RNA 的转基因小鼠品系培育 Mbnll 敲除小鼠来检查有毒 RNA 和结合蛋白之间的化学计量关系。其次，将检查肌盲蛋白通过与选择性剪接和/或其他因素相互作用来影响 CIC-1 氯离子通道水平的可能性。第三， 将测试使用重组腺相关病毒介导的野生型成体CIC-1表达可以挽救肌强直表型的假设。最后，我们将研究是否会因整个 Mbnl1 基因的缺失、组织特异性 Mbnl1 表达或所有三个肌盲基因（Mbnl1、Mbnl2/Mbnll、Mbnl3/Mblx）的组合丢失而导致其他与疾病相关的表型。