探究RYR2通过钙调磷酸酶信号通路调节心肌细胞成熟过程的作用和机制

心肌细胞成熟过程是形成健康成年心脏的关键步骤。钙处理能力是评价心肌细胞成熟度的关键指标之一，但是钙信号是否调节心肌细胞成熟并不清楚。申请人之前建立了腺相关病毒介导的基于CRISPR/Cas9的体细胞突变系统和心肌嵌合体分析方法，克服了在小鼠模型中研究心肌细胞成熟的技术瓶颈。利用这个新型实验平台，申请人发现钙通道RYR2（Ryanodine受体2）是心肌细胞成熟的关键调控因子。在此基础上，本项目将检验“RYR2通过增强钙调磷酸酶（CaN）信号促进心肌细胞成熟过程”这个科学假设。申请人将首先研究CaN信号对心肌细胞成熟的作用，然后探究RYR2对CaN活性的调节，最后尝试通过增强CaN信号修复RYR2缺失造成的心肌细胞成熟缺陷。由于钙信号在心脏病理过程中发挥关键作用，本项目不仅将阐明钙信号调节心肌细胞成熟的机制，而且可能以此为切入点，揭示钙信号通过调节心肌细胞成熟过程调节心脏病的新机制。

Cardiomyocyte maturation occurs in the last phase of heart development and is essential for the formation of the healthy heart in adults. Calcium handling capacity is a crucial measurement of cardiomyocyte maturity, but whether calcium signaling regulates cardiomyocyte maturation remains unclear. Previously, the applicant established adeno-associated virus-delivered CRISPR/Cas9-mediated somatic mutagenesis system and cardiac genetic mosaics analysis methods, which overcame the technical barriers in cardiomyocyte maturation research using mice as the model organism. Based on the new technical platform, the applicant discovered that RYR2, the crucial calcium channel that mediated cardiac calcium oscillation, played an essential role in cardiomyocyte maturation in a cell-autonomous manner. In this project, the applicant will further test the hypothesis that RYR2 promotes cardiomyocyte maturation by enhancing calcineurin signaling. The applicant will first determine the role of calcineurin signaling in cardiomyocyte maturation, and then examine the impact of RYR2 on calcineurin activity. Lastly, the applicant will test whether enhancing calcineurin activity could rescue the maturation defects in RYR2-depleted cardiomyocytes. Because calcineurin signaling also participates in cardiac pathogenesis, this project could not only uncover the mechanisms by which calcium signaling regulates cardiomyocyte maturation, but also potentially elucidate the contribution of cardiomyocyte maturation to cardiac pathogenesis.