RyR2-咖啡因结合位点的功能缺失W4645R突变:对同步性和心律失常发生的影响

IF 4.3 2区 生物学 Q2 CELL BIOLOGY
José-Carlos Fernández-Morales , Noemi Toth , Pinar Bayram , Taylor Rienzo , Martin Morad
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引用次数: 0

摘要

目的先前的研究发现,位于咖啡因结合位点的RyR2 W4645R突变与CPVT1病变有关。咖啡因与该位点的结合被认为会使羧基末端结构域与 Ca2+ 结合,从而使色氨酸残基(W4645)能够调节 RyR2 对 Ca2+ 的敏感性。为了深入了解RyR2 Ca2+结合的调控及其与咖啡因结合位点的相互作用,我们通过CRISPR/Cas9基因编辑技术在人类诱导多能干细胞衍生的心肌细胞(hiPSCCMs)中引入了W4645R-RyR2点突变,并将其与WT hiPSCCMs的Ca2+信号表型进行了比较。方法与结果W4645R-RyR2心肌细胞具有以下特征(1)ICa幅度或电压依赖性无明显变化;(2)CICR略有降低;(3)Ca2+-transients的松弛动力学改变,异丙肾上腺素敏感性无变化;(4)完全丧失咖啡因触发的Ca2+释放;(5)SR Ca2+泄漏增加,导致SR Ca2+含量降低40%,这是由心肌细胞对4-CmC的反应决定的;(6)钙火花和非同步自发SR Ca2+释放的发生率降低。结论W4645R-RyR2突变会导致咖啡因触发的SR Ca2+释放丧失,并增强SR Ca2+泄漏,从而导致非同步自发Ca2+释放,引发心律失常并损害心脏功能。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Loss-of-function W4645R mutation in the RyR2-caffeine binding site: implications for synchrony and arrhythmogenesis

Loss-of-function W4645R mutation in the RyR2-caffeine binding site: implications for synchrony and arrhythmogenesis

Aims

Previous studies have identified RyR2 W4645R mutation, located in the caffeine-binding site, to associate with CPVT1 pathology. Caffeine binding to its site is thought to displace the carboxyl-terminal domain to Ca2+-binding, allowing the tryptophan residue (W4645) to regulate Ca2+ sensitivity of RyR2. To gain insights into regulation of RyR2 Ca2+-binding and its interaction with caffeine-binding site, we introduced W4645R-RyR2 point mutation via CRISPR/Cas9 gene-editing in human induced pluripotent stem cell-derived cardiomyocytes (hiPSCCMs) and characterized their Ca2+-signaling phenotype compared to WT hiPSCCMs.

Methods and Results

W4645R-RyR2 cardiomyocytes had: (1) no significant change in ICa magnitude or voltage-dependence; (2) slightly reduced CICR; (3) altered relaxation kinetics of Ca2+-transients with no change in isoproterenol sensitivity; (4) complete loss of caffeine-triggered Ca2+ release; (5) larger SR Ca2+ leak resulting in 40 % lower SR Ca2+ content, as determined by myocytes’ response to 4-CmC; (6) lower incidence of calcium sparks and asynchronous spontaneous SR Ca2+ releases.

Conclusions

W4645R-RyR2 mutation induces loss of caffeine-triggered SR Ca2+ release and enhances SR Ca2+ leak that underlie asynchronous spontaneous Ca2+ releases, triggering arrhythmia and impairing cardiac function.

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来源期刊
Cell calcium
Cell calcium 生物-细胞生物学
CiteScore
8.70
自引率
5.00%
发文量
115
审稿时长
35 days
期刊介绍: Cell Calcium covers the field of calcium metabolism and signalling in living systems, from aspects including inorganic chemistry, physiology, molecular biology and pathology. Topic themes include: Roles of calcium in regulating cellular events such as apoptosis, necrosis and organelle remodelling Influence of calcium regulation in affecting health and disease outcomes
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