Novel cardioskeletal myopathy associated with MYL2

与 MYL2 相关的新型心脏骨骼肌病

• 批准号: 9272949
• 负责人: Danuta Szczesna-Cordary
• 金额: $ 49.88万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-06 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9272949
• 关键词: 3' Splice Site; Address; Affect; Age of Onset; Age-Months; Applications Grants; Arginine; Aspartic Acid; Benign; Binding Sites; Birth; C-terminal; Cardiac; Cardiac Death; Cardiomyopathies; Cessation of life; Complement; DNA; Disease; Electron Microscopy; Event; Familial Hypertrophic Cardiomyopathy; Family; Fiber; Foundations; Functional disorder; Genes; Genetic study; Glutamic Acid; Glutamine; Goals; Head; Heart; Heart Abnormalities; Heart Ventricle; Heart failure; Histopathology; Human; Hypertrophic Cardiomyopathy; Induced Mutation; Infant; Kinetics; Label; Link; Liquid Chromatography; Lysine; Malignant - descriptor; Measurement; Measures; Molecular; Mus; Muscle; Muscle Contraction; Muscle Fibers; Mutate; Mutation; Myocardial dysfunction; Myocardium; Myopathy; Myosin ATPase; Myosin Heavy Chains; Myosin Light Chain Kinase; Myosin Regulatory Light Chains; N-terminal; Nonmuscle Myosin Type IIA; Nonsense Codon; Papillary; Pathologic; Pathway interactions; Patients; Pattern; Phenotype; Phosphorylation; Point Mutation; Preparation; Proteomics; RNA Splicing; Recombinant Proteins; Regulation; Reporting; Risk; Sarcomeres; Severities; Signal Pathway; Site; Skeletal Muscle; Skeletal Muscle Myosins; Skin; Soleus Muscle; Splice-Site Mutation; Symptoms; Time; Transgenic Mice; Transgenic Organisms; Troponin; Valine; Variant; X ray diffraction analysis; X-Ray Diffraction; base; design; disease phenotype; experience; experimental study; in vivo; infancy; insight; mouse model; mutant; novel; papillary muscle; premature; public health relevance; reconstitution; retinal rods; skeletal; sudden cardiac death; tandem mass spectrometry; tool

 DESCRIPTION (provided by applicant): Infantile fiber-type I hypotrophy with simultaneously occurring severe onset of cardiomyopathy were previously reported in Dutch and Italian families and genetically linked to the MYL2 gene encoding for the human myosin regulatory light chain MLC2ventr/slow expressed in the ventricles and in slow-twitch skeletal muscles. Shortly after birth the patients experienced progressive slow-twitch skeletal myopathy and ultimately died of heart failure between 4 and 6 months of age. Dominant mutations in MYL2 have been known to cause familial hypertrophic cardiomyopathy (FHC) of extensive diversity in the course of the disease, age of onset and severity of symptoms. The mutation-specific dysregulation of the molecular events that trigger pathological remodeling of the heart, will be assessed using our transgenic (Tg) mice expressing the malignant: R58Q and D166V and benign: K104E mutations in MLC2ventr/slow. In addition to cardiac phenotypes, this application for the first time will include the slow-twitch skeletal muscle and the study of the splice site IVS6-1 mutation in MYL2 shown to cause severe myopathy in humans and premature death of IVS6-1-homozygous patients. AIM 1: Identify molecular mechanisms responsible for cardioskeletal dysfunction caused by MLC2ventr/slow mutations. We hypothesize that the mutation-induced structural changes trigger pathological remodeling of the heart and slow skeletal muscle leading to altered contractility and cardioskeletal myopathy. Proteomics study will be employed to identify the signaling pathways involved in cardioskeletal dysfunction associated FHC mutations. Structural phenotypes specific to MLC2ventr/slow mutations in the heart will be correlated with the respective phenotypes in the slow-twitch skeletal muscles using small angle X-ray diffraction patterns. Histopathology and electron microscopy (EM) will complement the effect of mutations on structural reorganization of the sarcomere in the heart and soleus muscle. Measurements of contractile force, force-pCa relationship and the myosin cross-bridge kinetics in skinned papillary/soleus muscle fibers from all proposed Tg mouse models of FHC will complete the phenotypic characterization of MLC2ventr/slow-specific cardioskeletal myopathy. Importantly, we will also study the IVS6-1 mutation associated with premature infantile cardiac death. AIM 2: Determine FHC induced cardiac phenotypes in vivo and explore novel rescue mechanisms in transgenic mice expressing constitutively phosphorylated P-MLC2. We hypothesize that by altering the Ca2+-dependent regulation of muscle contraction, D166V and R58Q mutations increase the propensity of affected patients toward malignant disease phenotypes. We also hypothesize that the underlying mechanisms relate to the steric inhibition of myosin light chain kinase dependent phosphorylation of MLC2. These hypotheses will be addressed using our recently developed double mutant Tg-S15D-D166V rescue mice, designed to mitigate the effects of the malignant D166V mutation with a constitutively phosphorylated Ser-15 (S15D).

 描述（由申请方提供）：先前在荷兰和意大利家族中报告了婴儿I型肌纤维营养不良伴心肌病重度发作，并与编码心室和慢收缩骨骼肌中表达的人肌球蛋白调节轻链MLC 2 ventr/slow的MYL 2基因遗传相关。出生后不久，患者出现进行性慢收缩骨骼肌病变，最终在4至6个月大时死于心力衰竭。已知MYL 2中的显性突变引起家族性肥厚型心肌病（FHC），其在疾病过程、发病年龄和症状严重程度方面具有广泛多样性。触发心脏病理性重塑的分子事件的突变特异性失调将使用在MLC 2 ventr/slow中表达恶性：R58 Q和D166 V以及良性：K104 E突变的转基因（Tg）小鼠来评估。除了心脏表型之外，该申请首次将包括慢收缩骨骼肌和MYL 2中剪接位点IVS 6 -1突变的研究，该突变显示会导致人类严重肌病和IVS 6 -1纯合子患者的过早死亡。目的1：确定MLC 2 ventr/slow突变引起的心脏骨骼功能障碍的分子机制。我们假设突变引起的结构变化触发心脏的病理性重塑，并减缓骨骼肌的收缩力改变和心脏骨骼肌病。蛋白质组学研究将用于确定与FHC突变相关的心脏骨骼功能障碍的信号通路。使用小角X射线衍射图，将心脏中MLC 2 ventr/slow突变特异性的结构表型与慢收缩骨骼肌中的相应表型相关联。组织学和电子显微镜（EM）将补充突变对心脏和比目鱼肌肌节结构重组的影响。测量来自所有拟定的FHC Tg小鼠模型的皮肤乳头肌/比目鱼肌纤维中的收缩力、力-pCa关系和肌球蛋白跨桥动力学将完成MLC 2 ventr/缓慢特异性心脏骨骼肌病的表型表征。重要的是，我们还将研究与早产婴儿心脏性死亡相关的IVS 6 -1突变。目标2：在体内确定FHC诱导的心脏表型，并在表达组成性磷酸化P-MLC 2的转基因小鼠中探索新的拯救机制。我们推测，通过改变肌肉收缩的Ca 2+依赖性调节，D166 V和R58 Q突变增加了受影响患者恶性疾病表型的倾向。我们还推测，潜在的机制涉及空间抑制肌球蛋白轻链激酶依赖的MLC 2磷酸化。这些假设将使用我们最近开发的双突变Tg-S15 D-D166 V拯救小鼠来解决，该小鼠旨在减轻具有组成性磷酸化Ser-15（S15 D）的恶性D166 V突变的影响。