RNA Dominance in Human Disease

描述(申请人提供)：虽然强直性肌营养不良症(DM)是成人中最常见的肌营养不良症，但这种神经肌肉疾病更严重的形式是先天性DM(COM)。虽然DMPK和ZNF9基因中不稳定的微卫星(CTG)n和(CCTG)n分别是由DMPK和ZNF9基因中不稳定的微卫星(CTG)n和(CCTG)n扩展引起的两种典型的成人型DM，但CDM只与DMPK基因中的&gt；1000个CTG重复数有关。这项拟议研究的长期目标是阐明导致CDM的分子事件，并利用这些信息开发这种疾病的新治疗策略。根据RNA显性模型，DM是一种RNA功能获得性疾病，其中突变的DMPK和ZNF9 RNA保留在细胞核中，在那里它们抑制肌盲样蛋白(MBNL)的功能。MBNL蛋白是一种选择性剪接因子，目前的证据表明，糖尿病是由新生儿蛋白亚型在成人体内滞留引起的。该提案的目的是同时使用基因敲除和转基因小鼠模型，将RNA优势模型扩展到CDM，并有四个实验目标。首先，将使用一种新的CDM转基因小鼠模型来验证特定的MBNL蛋白在胚胎发育过程中通过(CUG)n扩展进行隔离的假设。其次，CDM以新生儿低眼压和智力低下为特征，因此将研究MBNL蛋白在胚胎肌肉和脑发育过程中调节前mRNA加工的潜在作用。第三，CDM只与DMPK CTG重复扩增有关，以前的研究报告了CDM中DMPK以及相关基因DMWD和SIX5表达的变化。因此，将产生几个小鼠基因敲除系来检验这一假设，即CDM是由于MBNL和这些DMPK相关基因中的一个或几个基因的协调缺失造成的。第四，DMPK RNA的过度表达抑制了肌源性分化，因此将评估CDM是由MBNL和DMPK RNA结合蛋白共同隔离产生的假说。

DESCRIPTION (provided by applicant): Although myotonic dystrophy (DM) is the most prevalent muscular dystrophy in adults, a more severe form of this neuromuscular disease is congenital DM (COM). While the two characterized types of adult-onset DM, DM1 and DM2, are caused by unstable microsatellite (CTG)n and (CCTG)n expansions in the DMPK and ZNF9 genes, respectively, CDM is only associated with >1,000 CTG repeats in the DMPK gene. The longterm goals of the proposed research are to elucidate the molecular events which result in CDM and use this information to develop novel therapeutic strategies for this disease. According to the RNA dominance model, DM is an RNA gain-of-function disease in which mutant DMPK and ZNF9 RNAs are retained in the nucleus where they inhibit the functions of the muscleblind-like (MBNL) proteins. MBNL proteins are alternative splicing factors and current evidence suggests that DM disease is caused by the retention of neonatal protein isoforms in the adult. The objective of this proposal is to use both knockout and transgenic mouse models to extend the RNA dominance model to CDM using four experimental aims. First, the hypothesis that specific Mbnl proteins are sequestered during embryogenesis by (CUG)n expansions will be tested using a new transgenic mouse model for CDM. Second, CDM is characterized by neonatal hypotonia and mental retardation so potential roles for Mbnl proteins in the regulation of pre-mRNA processing during embryonic muscle and brain development will be investigated. Third, CDM is only associated with DMPK CTG repeat expansions and previous studies have reported changes in the expression of DMPK, as well as the linked genes DMWD and SIX5, in CDM. Therefore, several mouse knockout lines will be generated to test the hypothesis that CDM results from coordinate loss of MBNL and one, or several, of these DMPK associated genes. Fourth, overexpression of DMPK RNA inhibits myogenic differentiation so the hypothesis that CDM results from co-sequestration of MBNL and DMPK RNA-binding proteins will be evaluated.