Structure-Function of Calcium Channel Complexes in Cardiac Physiology and Disease

总结 心脏病是美国和世界范围内的主要死亡原因， 日益老龄化的人口。精确理解正常的分子机制 心脏生理学，以及它们在疾病中如何受损，对于确定新的药物靶点至关重要， 开发有效的新疗法来对抗心脏病。Ca 2+循环涉及局部信号传导， 表面L型Ca 2+（CaV1.2）通道和细胞内兰尼碱受体（RYR 2）负责Ca 2 +- 诱导的Ca 2+释放（CICR），是心脏兴奋-收缩偶联的基础。CaV1.2的调节异常 RYR 2导致异常钙信号传导，这是心脏病的不利标志。β-肾上腺素能 心肌收缩力的增强对战斗或逃跑反应至关重要，并通过CaV1.2增加介导 RYR 2的电流和致敏性;然而，在慢性应激下该途径的过度激活导致后 RYR 2通道的翻译修饰导致它们变得“渗漏”并导致心脏病理学， 以及CaV1.2的潜在有害亚细胞再分布。在知识方面存在着巨大的差距 关于CaV1.2和RYR 2在心脏中的功能组织和调节，无论是在疾病条件下; 他们的失调或功能障碍如何导致心脏病进展;以及他们是否以及如何能够 靶向有效治疗心力衰竭（HF）和其他心脏疾病。本计划项目补助金 (PPG)由四个项目和两个科学核心组成，旨在帮助解决这些关键问题。 差距。总体目标是确定CaV1.2和RYR 2调节局部Ca 2+信号传导的机制 在正常和衰竭的心脏中以前所未有的精确度。虽然每个项目都有自己的立足点， 这些研究都是创新和令人兴奋的，都依赖于核心小组提供的专业知识。 并通过项目间合作而得到丰富，而项目组合结构又极大地增强了项目间合作。所有四个项目 利用巴基斯坦基因组资源（PGR）（核心A），这是一个独特的个人队列，具有广泛的 HF和其他心脏病的表型和基因型数据以及高血缘关系率， 鉴定罕见截短突变纯合的个体（即，人类敲除）和其他 错义变体。此外，所有四个项目都涉及跨越单一基础研究的实验。 分子和细胞与动物模型（核心B;小鼠心脏生理学核心）。结合人类的误解/ 在PCR队列中发现的CaV1.2、RYR 2或关键调控蛋白的功能缺失突变， 结构-功能实验有望推进对基因型-表型关系的新理解， 涉及心脏中的Ca 2+循环蛋白的人类心血管疾病。我们预计拟议的研究将 对CaV1.2和RYR 2的结构-功能和调控产生新的见解，并促进其作为 心功能障碍的治疗靶点。四个项目的PI都有合作历史和跟踪 为CaV1.2和RYR 2分子生理学研究开发创新方法的记录。

SUMMARY Heart disease is the leading cause of death in the United States and worldwide, with a worsening trajectory due to increasingly aging populations. Precise understanding of the molecular mechanisms underlying normal cardiac physiology, and how they are compromised in disease, is critical for identifying new drug targets and developing effective new therapeutics to combat heart disease. Ca2+ cycling involving local signaling between surface L-type Ca2+ (CaV1.2) channels and intracellular ryanodine receptors (RYR2) is responsible for the Ca2+- induced Ca2+ release (CICR) that underlies cardiac excitation-contraction coupling. Dysregulation of both CaV1.2 and RYR2 contributes to abnormal calcium signaling that is an adverse hallmark of cardiac disease. β-adrenergic augmentation of cardiac contractility is crucial for the fight-or-flight response and is mediated by increased CaV1.2 current and sensitization of RYR2; yet, excessive activation of this pathway under chronic stress results in post- translational modifications of RYR2 channels that cause them to become ‘leaky’ and cause cardiac pathology, and also a potential harmful subcellular redistribution of CaV1.2. There are significant gaps in knowledge regarding CaV1.2 and RYR2 functional organization and regulation in heart under both and disease conditions; how their dysregulation or dysfunction contributes to heart disease progression; and whether and how they can be targeted for effective treatment of heart failure (HF) and other cardiac diseases. This Program Project Grant (PPG) comprises four Projects and two Scientific Cores that have been put together to help address these critical gaps. The overarching goal is to define the mechanisms that regulate local Ca2+ signaling by CaV1.2 and RYR2 in normal and failing hearts with unprecedented precision. While each project stands on its own footing as far as being comprised of innovative and exciting research, all are dependent on the expertise provided by the Cores and are enriched by interproject collaborations that are greatly enhanced by the PPG structure. All four Projects leverage the Pakistan Genome Resource (PGR) (Core A), a unique cohort of individuals with extensive phenotype and genotype data on HF and other cardiac diseases and high rates of consanguinity enabling identification of individuals homozygous for rare truncating mutations (i.e., human knockouts) and other missense variants. Moreover, all four Projects involve experiments that span fundamental studies on single molecules and cells to animal models (Core B; Mouse Cardiac Physiology Core). Combining human missense / loss of function mutations found in the PGR cohort in CaV1.2, RYR2 or key regulatory proteins with in-depth structure-function experiments promises to advance new understanding of genotype-phenotype relationships in human cardiovascular diseases involving Ca2+ cycling proteins in the heart. We expect the proposed studies to yield new insights into structure-function and regulation of CaV1.2 and RYR2 and advance their utility as therapeutic targets for cardiac dysfunction. The PIs of the four projects have a collaboration history and track record of developing innovative approaches for studies of CaV1.2 and RYR2 molecular physiology.