Myogenesis and RNA Biogenesis in a Mouse Model of OPMD

OPMD 小鼠模型中的肌发生和 RNA 生物发生

• 批准号: 9050130
• 负责人: Katherine Elizabeth Vest
• 金额: $ 5.61万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-02-22 至 2019-02-21
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9050130
• 关键词: Acute; Adult; Affect; Age-Months; Alanine; Alleles; Animal Model; Animals; Architecture; Aspiration Pneumonia; Automobile Driving; Binding Proteins; Biogenesis; Biological Assay; Bromodeoxyuridine; Cell Nucleus; Cell physiology; Cells; Cellular biology; Cessation of life; Choking; Defect; Deglutition; Deglutition Disorders; Development; Disease; Enteral Feeding; Event; Eyelid structure; Future; Gastrocnemius Muscle; Genes; Genotype; Goals; In Vitro; Individual; Injury; Knock-in Mouse; Label; Laboratories; Late-Onset Disorder; Length; Limb structure; Measures; Messenger RNA; Metabolism; Mitotic; Modeling; Molecular; Muscle; Muscle satellite cell; Mutation; Myoblasts; Myopathy; N-terminal; Natural regeneration; Nuclear; Oculopharyngeal Muscular Dystrophy; Operative Surgical Procedures; Pathology; Pathway interactions; Patients; Pharyngeal structure; Phenotype; Play; Poly A; Poly(A) Tail; Polyadenylation; Process; Proliferating; Proteins; Quality of life; RNA; RNA Processing; Reporting; Research; Resistance; Risk; Signal Transduction; Skeletal Muscle; Specificity; Staging; Stem cells; Stretching; Symptoms; System; Time; Tissues; Transcript; Wild Type Mouse; Work; age related; base; cell type; experience; improved; in vivo; innovation; mouse model; muscle regeneration; mutant; myogenesis; novel; pharynx muscle; polyalanine; programs; promoter; public health relevance; regenerative; research study; satellite cell; therapeutic target

 DESCRIPTION (provided by applicant): Oculopharyngeal muscular dystrophy (OPMD) is a late onset disorder causing weakness of eyelid and pharyngeal, and proximal limb muscles. A major symptom of OPMD is swallowing difficulty (dysphagia) due to weakened pharyngeal muscles. Patients often experience choking or aspiration pneumonia, and may require a feeding tube in later stages of disease. At present, the exact molecular mechanisms of pathology are unknown. The only available options to treat dysphagia in OPMD patients are temporary, usually surgical procedures to decrease resistance in the throat to improve swallowing. Autosomal dominant OPMD patients harbor a mutation in the gene for the nuclear mRNA processing protein PABPN1, which causes the N-terminal polyalanine stretch of PABPN1 to expand from 10 to 11-18 residues. PABPN1 plays key roles in mRNA processing including polyadenylation of nuclear transcripts and the alternative polyadenylation and cleavage pathway. PABPN1 is expressed in all tissues, but mutation causes late-onset pathology only in a specific subset of skeletal muscles. How mutation of this ubiquitously expressed protein leads to late-onset pathology in specific tissues remains an open question. Skeletal muscle tissue is capable of regeneration. Closely associated with multinucleated, post-mitotic myofibers in adult muscles is a pool of stem cells known as satellite cells. Upon muscle damage, satellite cells become activated to proliferate, differentiate, and fuse with each other and existing myofibers. This process, known as myogenesis, restores muscle architecture and function. Interestingly, some of the muscles most severely affected by OPMD, including pharyngeal muscles, are more highly regenerative than other skeletal muscles. The hypothesis driving the work proposed here is that mutant PABPN1 disrupts mRNA biogenesis and decreases the myogenic capacity of pharyngeal satellite cells, leading to pathology due to the unusually high demand for myogenesis in pharyngeal muscles. We will take an integrative in vitro and in vivo approach using a novel mouse model that is the first to accurately copy the genotype of OPMD patients. This mouse model expresses mutant PABPN1 in all tissues. Because PABPN1 is known to impact mRNA metabolism, satellite cells from pharyngeal muscles will be isolated and assayed for RNA phenotypes (Aim 1). The myogenic program can be recapitulated in vitro and used to quantify specific stages of myogenesis. Therefore, primary activated satellite cells, known as myoblasts, from pharyngeal muscles will be isolated and used to quantify the effects of mutant PABPN1 on the specific stages of myogenesis in vitro (Aim 2). This model will then be used to assay the effect of mutant PABPN1 on myogenesis in pharyngeal muscle in vivo (Aim 3). The long-term objective of this work is to understand how mutant PABPN1 affects pharyngeal satellite cell biology and potentially identify targets for new pharmacologic OPMD treatments.

 描述（由适用提供）：咽咽肌营养不良（OPMD）是一种晚期发作障碍，导致眼睑和咽部肌肉弱，以及近端的肢体肌肉。 OPMD的主要症状是由于咽部肌肉减弱而引起的困难（吞咽困难）。患者经常会出现窒息或抽吸肺炎，可能需要在疾病的后期喂养管。目前，治疗OPMD患者吞咽困难的唯一可用选择的确切分子机制是暂时的，通常是手术程序，可以降低喉咙的耐药性以改善吞咽。常染色体显性OPMD患者具有核mRNA加工蛋白PABPN1的基因中的突变，这会导致PABPN1的N末端多酰氨酸延伸从10个残留物膨胀到11-18个残留物。 PABPN1在mRNA加工中起关键作用，包括核转录物的聚腺苷酸化以及替代的聚腺苷酸化和切割途径。 PABPN1在所有组织中均表达，但是突变仅在骨骼肌的特定子集中引起晚期病理。这种无处不在表达的蛋白质的突变如何导致特定组织中迟到的病理仍然是一个开放的问题。骨骼肌组织能够再生。与成年肌肉中的多核后有丝质肌纤维密切相关的是一种称为卫星细胞的干细胞。肌肉损伤后，卫星细胞被激活以增殖，分化和融合彼此和现有肌纤维。这个被称为肌发生的过程可恢复肌肉结构和功能。有趣的是，某些受OPMD影响最严重的肌肉（包括咽肌肉）比其他骨骼肌更具再生性。推动此处提出的工作的假设是，突变体PABPN1破坏了mRNA生物发生并降低了咽卫星细胞的肌原性能力，导致病理学导致病理学，这是由于对咽肌的肌发生异常高的需求。我们将使用一种新型小鼠模型进行体外和体内综合方法，该模型是第一个准确复制OPMD患者基因型的模型。该小鼠模型在所有组织中表达突变的PABPN1。由于已知PABPN1会影响mRNA代谢，因此将分离出来自咽部肌肉的卫星细胞并分配为RNA表型（AIM 1）。可以在体外概括肌源程序，并用于量化肌发生的特定阶段。因此，将分离出来自咽部肌肉的原代活化的卫星细胞，称为肌细胞，并用于量化突变体Pabpn1在体外特定阶段的影响（AIM 2）。然后，该模型将用于断言突变体PABPN1对体内咽肌的肌发生的影响（AIM 3）。这项工作的长期目标是了解突变体PABPN1如何影响咽卫星细胞生物学，并有可能识别新药理OPMD治疗的靶标。