Translational Control of Cardiac Excitability

项目概要/摘要 心脏的规律跳动需要特定离子通道群的精心策划的表达 心室复极、起搏、传导等功能。最近的进展建立了一个新的 协调渠道表达的方式。该提案的中心假设是 相互作用的 mRNA 物种的“微翻译组”编码功能相关的蛋白质，例如 产生心室动作电位。这些离子通道共翻译组装成大分子 控制高阶心脏兴奋性的复合物。该提案的目的是解决以下问题的组成部分： 使用单分子荧光原位介导心脏复极的共翻译复合物 在胚胎 (iPSC) 和成人中进行杂交结合蛋白质免疫荧光 (smFISH/IF) 心肌细胞。关于这些物质的组成、化学计量和细胞定位的假设 将测试复合物。这些复合物是否调节其他心脏功能，例如起搏，将 被确定。介导 mRNA 关联的机制，例如候选 RNA 结合蛋白 (RBP) 通过亲和捕获质谱鉴定以及直接 mRNA 相互作用将得到解决。这些 预计研究将阐明控制心脏兴奋性的新机制并确定新的机制 疾病和治疗开发的目标。

PROJECT SUMMARY/ABSTRACT The regular beating of the heart requires the orchestrated expression of ion channel ensembles specific for ventricular repolarization, pacemaking, conduction and other functions. Recent progress establishes a new way in which channel expression is coordinated. The central hypothesis of this proposal is that a “microtranslatome” of interacting mRNA species encodes functionally related proteins, such as those producing the ventricular action potential. These ion channels assemble cotranslationally into macromolecular complexes that govern higher-order cardiac excitability. The aims of the proposal are to resolve components of cotranslational complexes mediating cardiac repolarization using single-molecule fluorescence in situ hybridization combined with protein immunofluorescence (smFISH/IF) in both embryonic (iPSC) and adult cardiomyocytes. Hypotheses regarding the composition, stoichiometry and cellular localization of these complexes will be tested. Whether such complexes regulate other cardiac functions, such as pacemaking, will be determined. Mechanisms mediating mRNA association, such as candidate RNA binding proteins (RBPs) identified with affinity-capture mass spectrometry, and direct mRNA interactions, will be resolved. These studies are expected to illuminate new mechanisms by which cardiac excitability is controlled and identify novel targets for disease and therapeutic development.