Myotonic dystrophy type 1 (DM1) is a multisystem disorder resulting from the expansion of a CTG repeat in the 3’-untranslated region of DMPK gene. It has been proposed that CTG expansion not only affects the expression of the DMPK gene, but also alters the expression of its neighboring genes, SIX5 and DMWD. Moreover, RNA disease model suggest that mutant DMPK mRNAs are retained in the nucleus and sequester muscleblind-like (MBNL) proteins. Mouse models carrying mutations in DMPK, SIX5 or MBNL1 have been generated; however, none of them faithfully recapitulates the multisystemic and often severe phenotype seen in human patients. We and others thus hypothesize that CTG expansion may simultaneously repress multiple gene expression. To test this hypothesis, in this study, we will adopt haploid cell-mediated semi-cloned technology and generate haploid cells carrying compound mutations of DMPK, SIX5, MBNL1 or DMWD. By injection of mutant haploid cells into MII oocytes, semi-cloned (SC) mice carrying heterozygous mutant genes can be efficiently produced. We will then perform phenotype analysis to ensure that one mouse model with combined mutations can fully mimic DM1 phenotypes. We believe that, from this proposed study, we will not only reveal the underlying mechanism of DM1 through establishment of an ideal mouse model, but also provide a new system for study of molecular mechanism of complex diseases.
1型强直性肌营养不良症(DM1)是最常见的肌肉萎缩症,会出现多系统病变。该病是由于在强直性肌营养不良蛋白激酶(DMPK) 基因的3’末端出现了CTG三核苷酸的异常扩增所导致的,但是针对CTG异常扩增产生病变的分子机制还不清楚。已有的假说显示,DMPK本身,DMPK相邻基因包括SIX5和DMWD,或者MBNL1表达异常是发病的关键原因。然而,这些基因的单独敲除小鼠却不能模拟DM1的所有症状。因此,我们认为,DM1是CTG重复引起多个基因表达异常所致的。为了验证这一假设,本研究中,我们利用了孤雄单倍体干细胞介导的半克隆技术,在单倍体细胞中针对DMPK、SXI5、DMWD和MBNL1进行不同组合敲除,然后通过注入卵母细胞中一步获取携带不同组合突变的杂合小鼠,进而分析表型,以期获得能模拟DM1的理想小鼠模型。本研究将揭示DM1的发病机制,建立理想的DM1小鼠模型,从而为研究疾病发生提供新的手段。
1型强直性肌营养不良症(DM1)是最常见的肌肉萎缩症,会出现多系统病变。该病是由于在强直性肌营养不良蛋白激酶(DMPK) 基因的3’末端出现了CTG三核苷酸的异常扩增所导致的,但是针对CTG异常扩增产生病变的分子机制还不清楚。已有报道提示DMPK、DMPK相邻基因包括SIX5和DMWD,或者MBNL1表达异常是发病的关键原因。然而,这些基因的单独敲除小鼠却不能模拟DM1的所有症状。因此,我们假设DM1是多基因异常突变的结果。本项目利用了孤雄单倍体干细胞介导的半克隆技术,在单倍体细胞中针对DMPK、SXI5、DMWD和MBNL1进行不同组合敲除,然后通过注入卵母细胞中一步获取携带不同组合突变的杂合小鼠,进而分析表型。我们发现四基因敲除小鼠能模拟几乎所有的DM1症状,是理想的DM1疾病小鼠模型,并揭示了DM1的发病机制,发现了能够明显改善DM1肌肉干细胞功能的小分子化合物。该模型的建立为研究疾病发生提供新的手段,为研究DM1分子机制并进行药物筛选奠定了基础。该项目的实施为构建携带多基因突变小鼠模型提供了一个快捷便利的实验思路和方案,也为研究多基因疾病表型机理提供了可能性。
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数据更新时间:2023-05-31
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