Molecular Pathogenesis of Myotonic Dystrophy

描述（由申请人提供）：强直性肌营养不良（1型）（DM 1）是成人中最常见的肌营养不良形式，由DMPK基因3'非翻译区的CTG扩增引起。DM 1的发病机制是由于含有扩增的CUG重复序列（CUGexp RNA）的DMPK mRNA的功能获得。CUGexp RNA改变RNA结合蛋白的功能，例如肌盲样（MBNL）和CUGBP，Elav样家族（CELF）的成员。MBNL 1被隔离在扩展的CUG重复序列上，导致功能丧失，而CELF 1由于蛋白激酶C（PKC）的激活而上调，导致其磷酸化和稳定化。虽然关于CUGexp RNA表达的分子后果已经了解了很多，但CUGexp RNA引起进行性肌肉萎缩的机制仍然未知。为了研究肌肉中的致病机制，我们建立了肌肉特异性四环素诱导的转基因小鼠模型，表达含有960个CUG重复的DMPK mRNA的最后1200个核苷酸。诱导表达CUGexp RNA的动物发生进行性和严重的组织病理学和肌肉萎缩。我们将使用该模型来确定年龄在疾病进展中的作用，并确定去除CUGexp RNA后是否会消退的慢性消耗的特征。我们将在小鼠模型中使用遗传方法来确定CELF 1的丢失或MBNL 1和/或MBNL 2的获得是否阻止或挽救肌肉表型。我们将确定PKC通路是否在DM 1骨骼肌中被激活，如在DM 1心脏中所示，以及PKC抑制是否挽救肌肉表型。来自DM 1小鼠模型和DM 1肌肉样品的肌肉的RNA-seq数据将用于鉴定，然后测试关键异常基因表达和可变剪接事件在肌肉病理学中的作用。我们还将进行研究，以确定CELF/MBNL独立的肌肉病理机制。本研究的目的是了解CUGexp RNA导致进行性肌肉损失的分子机制，从而增加潜在治疗靶点的数量。

DESCRIPTION (provided by applicant): Myotonic dystrophy (type 1) (DM1) is the most prevalent form of muscular dystrophy in adults and is caused by a CTG expansion in the 3' untranslated region of the DMPK gene. Pathogenesis in DM1 is due to a gain of function of the DMPK mRNA that contains expanded CUG repeats (CUGexp RNA). The CUGexp RNA alters the functions of RNA binding proteins such as members of the muscleblind like (MBNL) and CUGBP, Elav-like family (CELF) families. MBNL1 is sequestered on the expanded CUG repeats producing a loss of function while CELF1 is up regulated due to activation of protein kinase C (PKC) leading to its phosphorylation and stabilization. While much has been learned regarding the molecular consequences of CUGexp RNA expression, the mechanisms by which CUGexp RNA causes progressive muscle wasting remains unknown. To study the pathogenic mechanism in muscle, we developed a muscle specific tetracycline inducible transgenic mouse model expressing the last 1200 nucleotides of the DMPK mRNA containing 960 CUG repeats. Animals induced to express the CUGexp RNA develop progressive and severe histopathology and muscle wasting. We will use this model to determine the role of age in disease progression and identify the features of chronic wasting that do and do not regress upon removal of the CUGexp RNA. We will use genetic approaches in the mouse model to determine whether the loss of CELF1 or gain of MBNL1 and/or MBNL2 prevents or rescues the muscle phenotype. We will determine whether the PKC pathway is activated in DM1 skeletal muscle, as has been shown in DM1 heart, and whether PKC inhibition rescues the muscle phenotype. RNA-seq data from muscle of DM1 mouse models and DM1 muscle samples will be used to identify and then test the roles of critical aberrant gene expression and alternative splicing events in muscle pathology. We will also perform studies to identify CELF/MBNL-independent mechanisms of muscle pathology. The goal of this study is to develop an understanding of the molecular mechanisms by which CUGexp RNA leads to progressive muscle loss and thereby increase the number of potential therapeutic targets.