The molecular mechanisms underlying arrhythmogenic right ventricular dysplasia/cardiomyopathy

致心律失常性右心室发育不良/心肌病的分子机制

• 批准号: 8884263
• 负责人: Farah Sheikh
• 金额: $ 38.75万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8884263
• 关键词: Accounting; Adult; Affect; Arrhythmogenic Right Ventricular Dysplasia; Autophagocytosis; Autophagosome; Behavior; Binding; Biochemical; Biological Models; Biology; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cells; Clinical; Complex; Congenital Heart Defects; Connexins; DNA Sequence Alteration; Data; Defect; Desmosomes; Diagnosis; Disease; Employee Strikes; Event; Exhibits; Family; Funding; Generations; Genes; Genetic; Genetic Models; Goals; Grant; Health; Heart; Heart Diseases; Human; In Vitro; Inherited; Intercellular Junctions; Knockout Mice; Link; Lysosomes; Mediating; Microscopy; Modeling; Molecular; Molecular Genetics; Morphology; Mus; Mutation; Myocardium; Patients; Physiological; Physiology; Protein Family; Proteins; Role; SNAP receptor; Structure; Sudden Death; Testing; Validation; Yeasts; base; desmoplakin; effective therapy; human tissue; in vivo; induced pluripotent stem cell; insight; lentiviral-mediated; loss of function; member; mouse model; novel; overexpression; protein degradation; protein function; public health relevance; small hairpin RNA; soluble NSF attachment protein; sudden cardiac death; targeted treatment; tool; yeast two hybrid system

 DESCRIPTION (provided by applicant): This is an application for a competitive renewal of our grant R01 HL095780-01. We were originally funded to investigate mechanisms underlying the role of the known central desmosomal component, desmoplakin, in the clinical and cellular features associated with the genetic-based heart disease, arrhythmogenic right ventricular cardiomyopathy (ARVC) that causes sudden death in the young, by generating and characterizing novel desmoplakin deficient model systems. For this application, we uncovered synaptosomal associated protein 29 (SNAP29) as a novel desmoplakin associated protein in the adult human heart in a yeast-two hybrid screen that we show has relevance to ARVC, by leveraging our novel genetic mouse and human cardiac models of ARVC. Although the role of SNAP29 is unknown in the heart, we show that SNAP29 co-localizes to cardiac muscle desmosomal cell-cell junctions in the adult mouse and human heart as well as human induced pluripotent stem cell derived cardiac cells (hiPSC). Furthermore, SNAP29 localization and/or levels at desmosomal junctions are lost in hearts from our mouse model of ARVC and ARVC hiPSC-derived cardiac cells that exhibit striking desmosomal defects and arrhythmogenic behavior. Generation of novel SNAP29 deficient mouse models (global and cardiomyocyte-specific) revealed striking cardiac defects including (i) autophagic defects at the cardiac muscle cell junction (accumulation of autophagic/lysosomal markers and structures at the cell junction) that specifically impacted desmosomal protein levels and (ii) cardiac morphology defects. Data from our mouse model of ARVC provides validation to this mechanism as we reveal that their hearts exhibit similar defects in autophagic control at the cardiac muscle cell junction. We hypothesize that SNAP29 regulates desmosomal protein levels and function in cardiac muscle by controlling desmosomal turnover via autophagy-mediated mechanisms and its loss will trigger loss of desmosomal protein levels as well as function and ultimately cause ARVC. We aim to determine: (i) the functional role of SNAP29 in the heart by characterizing SNAP29 loss of function mouse models, (ii) the relevance of SNAP29-DSP interaction in human ARVC and cardiomyocytes by expressing human ARVC mutations and using hiPSCs as a tool, and (iii) the SNAP29-dependent events in autophagy that control desmosomal levels/turnover, by analyzing defects in cardiac autophagy (induction and flux) and relevant desmosome targets using SNAP29 loss of function models and overexpression of SNAP29 in an in vitro ARVC model.

 描述（由申请人提供）：这是我们的 R01 HL095780-01 拨款的竞争性更新申请。我们最初的资助是为了研究已知的中央桥粒成分桥粒蛋白在与相关临床和细胞特征相关的作用机制。通过生成和表征新型桥粒斑蛋白缺陷模型，研究导致年轻人猝死的遗传性心脏病、致心律失常性右心室心肌病 (ARVC)对于此应用系统，我们通过利用我们的新型基因小鼠和人类心脏，在酵母-两种杂交筛选中发现了突触体相关蛋白 29 (SNAP29) 作为成人心脏中的一种新型桥粒斑蛋白相关蛋白，我们证明该蛋白与 ARVC 相关。尽管 SNAP29 在心脏中的作用尚不清楚，但我们发现 SNAP29 在成年小鼠和人类心脏中共定位于心肌桥粒细胞-细胞连接处。此外，在我们的 ARVC 小鼠模型和 ARVC hiPSC 衍生心脏细胞中，SNAP29 在桥粒连接处的定位和/或水平丢失，这些细胞表现出明显的桥粒缺陷和致心律失常行为。新型 SNAP29 缺陷小鼠模型（整体和心肌细胞特异性）的生成揭示了显着的心脏缺陷，包括 (i) 心肌细胞连接处的自噬缺陷（细胞连接处自噬/溶酶体标记物和结构的积累）特别影响桥粒蛋白水平和（ii）心脏形态缺陷，来自我们的 ARVC 小鼠模型的数据验证了这一机制，因为我们揭示了它们的心脏在我们研究发现，SNAP29 通过自噬介导的机制控制桥粒周转，从而调节心肌中的桥粒蛋白水平和功能，其丢失将引发桥粒损失。我们的目标是通过表征 SNAP29 功能丧失小鼠模型来确定 SNAP29 在心脏中的功能作用，(ii) SNAP29-DSP 相互作用在人类 ARVC 和 ARVC 中的相关性。通过表达人类 ARVC 突变并使用 hiPSC 作为工具来研究心肌细胞，以及 (iii) 通过分析心脏缺陷来控制桥粒水平/周转的自噬中的 SNAP29 依赖性事件使用 SNAP29 功能丧失模型和体外 ARVC 模型中 SNAP29 的过度表达来研究自噬（诱导和通量）和相关桥粒靶标。