Investigating mechanisms of skeletal muscle degeneration in Myotonic Dystrophy

研究强直性肌营养不良骨骼肌变性的机制

• 批准号: 7574524
• 负责人: James P Orengo
• 金额: $ 3.62万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7574524
• 关键词: 3' Untranslated Regions; Address; Adult; Alternative Splicing; Biological Assay; Breeding; CD34 gene; Cause of Death; Cell Culture Techniques; Cell Nucleus; Cerebellar Ataxia; Characteristics; Control Animal; Defect; Degenerative Disorder; Development; Disease; Disease Progression; Electron Microscopy; Electrophoresis; Event; Exercise; Exhibits; Fragile X Syndrome; Friedreich Ataxia; Genes; Genetic Recombination; Genetic Transcription; Goals; Heart; Histology; Huntington Disease; IGF1 gene; Individual; Injury; Insulin Resistance; Investigation; Lead; Magnetic Resonance Imaging; Measures; Mediating; Modeling; Molecular; Mus; Muscle; Muscle Cells; Muscle Fibers; Muscle function; Muscle satellite cell; Muscular Atrophy; Muscular Dystrophies; Myf-6 myogenic factor; Myogenin; Myotonia; Myotonic Dystrophy; Natural regeneration; Nuclear; Nucleotides; PAX7 gene; Pathology; Pathway interactions; Patients; Pattern; Phenotype; Phosphorylation; Play; Proliferating; Protein Kinase C Alpha; Proteins; RNA; RNA Splicing; Regulation; Reverse Transcriptase Polymerase Chain Reaction; Role; Skeletal Muscle; Spinal; Symptoms; Tamoxifen; Testing; Tetanus Helper Peptide; Time; Tissues; Toxic effect; Transgenes; Transgenic Animals; Transgenic Mice; Transgenic Organisms; Untranslated Regions; Western Blotting; Work; in vivo; injured; insight; mouse model; muscle degeneration; muscle regeneration; novel; overexpression; postnatal; regenerative; repaired; response; satellite cell; wasting

DESCRIPTION (provided by applicant): Myotonic dystrophy (DM) is the most common cause of adult-onset muscular dystrophy. This multi-systemic disorder is marked by a debilitating and progressive loss of skeletal muscle function. The major form of DM is caused by expanded CTG repeats located in the 3' UTR of the DMPK gene. It is now clear that after transcription the expanded repeats remain trapped in the nucleus and lead to a trans-dominant misregulation of alternative splicing. Although many symptoms in DM, such as myotonia and insulin resistance, are explained by particular misregulated splicing events, the mechanism by which skeletal muscle degeneration arises remains unknown. This proposal will investigate the cause of muscle degeneration using a novel transgenic mouse model generated in the sponsor's lab. In Specific Aim 1 two mouse lines will be bred to produce bi-transgenic animals which upon tamoxifen induction express 960 CUG repeats specifically in skeletal muscle. Using a number of cellular, molecular and functional assays bitransgenic mice will be tested to see how well they recapitulate the DM phenotype. In Specific Aim 2, the role of defective satellite cells in DM muscle degeneration will be examined. Satellite cells are a subpopulation of muscle cells which proliferate, differentiate and fuse with existing myotubes in order to repair injured muscle. Exercise, muscle damage and degenerative disorders are known to activate satellites cells; however in DM they fail to be activated. Satellite cells will be isolated from our bi-transgenic mice, and mechanisms leading to defective activation will be further studied. In Specific Aim 3 a splicing factor, CUGBP1, which regulates developmental alternative splicing patterns in post-natal development, will be investigated. Although CUGBP1 levels decrease in adulthood, they remain unexpectedly high in DM patients and correlate with misregulation of certain alternative splicing events. CUGBP1 steady state levels will be assayed in bi-transgenic mice after tamoxifen treatment. A potential mechanism for increased CUGBP1 protein levels via phosphorylation through the PKC alpha pathway will also be examined. The proposed work aims to elucidate the pathogenic mechanisms responsible for skeletal muscle degeneration in the leading cause of adult-onset muscular dystrophy, Myotonic Dystrophy. Information gained from this project may also provide insight into other tri-nucleotide expansion repeat disorders, such as Huntington's Chorea, Friedreich's Ataxia, Spinal Cerebellar Ataxia and Fragile X syndrome.

描述（由申请人提供）： 强直性肌营养不良（DM）是成人型肌营养不良症最常见的原因。这种多系统疾病的特点是骨骼肌功能的衰弱和进行性丧失。DM的主要形式是由位于DMPK基因的3' UTR中的扩增的CTG重复序列引起的。现在清楚的是，转录后，扩展的重复序列仍然被困在细胞核中，并导致选择性剪接的反式显性失调。虽然糖尿病的许多症状，如肌强直和胰岛素抵抗，是由特定的错误调节剪接事件解释的，骨骼肌变性发生的机制仍然未知。本提案将使用申办方实验室生成的新型转基因小鼠模型研究肌肉变性的原因。在特定目标1中，将培育两个小鼠品系以产生双转基因动物，其在他莫昔芬诱导后在骨骼肌中特异性表达960个CUG重复。将使用许多细胞、分子和功能测定来测试双转基因小鼠，以观察它们再现DM表型的程度。在具体目标2中，将检查缺陷卫星细胞在DM肌肉变性中的作用。卫星细胞是肌细胞的亚群，其增殖、分化并与现有的肌管融合以修复受损的肌肉。已知运动、肌肉损伤和退行性疾病会激活卫星细胞;然而，在DM中，它们不能被激活。卫星细胞将从我们的双转基因小鼠中分离出来，导致缺陷激活的机制将进一步研究。在具体目标3剪接因子，CUGBP 1，它调节发育的选择性剪接模式在出生后的发展，将进行调查。虽然CUGBP 1水平在成年期下降，但在DM患者中仍然出乎意料地高，并与某些选择性剪接事件的误调节相关。将在他莫昔芬处理后的双转基因小鼠中测定CUGBP 1稳态水平。还将研究通过PKC α途径磷酸化增加CUGBP 1蛋白水平的潜在机制。这项工作的目的是阐明骨骼肌变性的致病机制，在成人发病的肌营养不良症，强直性肌营养不良症的主要原因。从这个项目中获得的信息也可以提供对其他三核苷酸扩展重复障碍的深入了解，如亨廷顿舞蹈病，弗里德赖希共济失调，脊髓小脑共济失调和脆性X综合征。