José-Carlos Fernández-Morales , Noemi Toth , Pinar Bayram , Taylor Rienzo , Martin Morad
{"title":"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","authors":"José-Carlos Fernández-Morales , Noemi Toth , Pinar Bayram , Taylor Rienzo , Martin Morad","doi":"10.1016/j.ceca.2024.102925","DOIUrl":null,"url":null,"abstract":"<div><h3>Aims</h3><p>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 Ca<sup>2+</sup>-binding, allowing the tryptophan residue (W4645) to regulate Ca<sup>2+</sup> sensitivity of RyR2. To gain insights into regulation of RyR2 Ca<sup>2+</sup>-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 (hiPSC<img>CMs) and characterized their Ca<sup>2+</sup>-signaling phenotype compared to WT hiPSC<img>CMs.</p></div><div><h3>Methods and Results</h3><p>W4645R-RyR2 cardiomyocytes had: <strong>(1)</strong> no significant change in I<sub>Ca</sub> magnitude or voltage-dependence; <strong>(2)</strong> slightly reduced CICR; <strong>(3)</strong> altered relaxation kinetics of Ca<sup>2+</sup>-transients with no change in isoproterenol sensitivity; <strong>(4)</strong> complete loss of caffeine-triggered Ca<sup>2+</sup> release; (<strong>5</strong>) larger SR Ca<sup>2+</sup> leak resulting in 40 % lower SR Ca<sup>2+</sup> content, as determined by myocytes’ response to 4-CmC; <strong>(6)</strong> lower incidence of calcium sparks and asynchronous spontaneous SR Ca<sup>2+</sup> releases.</p></div><div><h3>Conclusions</h3><p>W4645R-RyR2 mutation induces loss of caffeine-triggered SR Ca<sup>2+</sup> release and enhances SR Ca<sup>2+</sup> leak that underlie asynchronous spontaneous Ca<sup>2+</sup> releases, triggering arrhythmia and impairing cardiac function.</p></div>","PeriodicalId":9678,"journal":{"name":"Cell calcium","volume":null,"pages":null},"PeriodicalIF":4.3000,"publicationDate":"2024-06-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Loss-of-function W4645R mutation in the RyR2-caffeine binding site: implications for synchrony and arrhythmogenesis\",\"authors\":\"José-Carlos Fernández-Morales , Noemi Toth , Pinar Bayram , Taylor Rienzo , Martin Morad\",\"doi\":\"10.1016/j.ceca.2024.102925\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Aims</h3><p>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 Ca<sup>2+</sup>-binding, allowing the tryptophan residue (W4645) to regulate Ca<sup>2+</sup> sensitivity of RyR2. To gain insights into regulation of RyR2 Ca<sup>2+</sup>-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 (hiPSC<img>CMs) and characterized their Ca<sup>2+</sup>-signaling phenotype compared to WT hiPSC<img>CMs.</p></div><div><h3>Methods and Results</h3><p>W4645R-RyR2 cardiomyocytes had: <strong>(1)</strong> no significant change in I<sub>Ca</sub> magnitude or voltage-dependence; <strong>(2)</strong> slightly reduced CICR; <strong>(3)</strong> altered relaxation kinetics of Ca<sup>2+</sup>-transients with no change in isoproterenol sensitivity; <strong>(4)</strong> complete loss of caffeine-triggered Ca<sup>2+</sup> release; (<strong>5</strong>) larger SR Ca<sup>2+</sup> leak resulting in 40 % lower SR Ca<sup>2+</sup> content, as determined by myocytes’ response to 4-CmC; <strong>(6)</strong> lower incidence of calcium sparks and asynchronous spontaneous SR Ca<sup>2+</sup> releases.</p></div><div><h3>Conclusions</h3><p>W4645R-RyR2 mutation induces loss of caffeine-triggered SR Ca<sup>2+</sup> release and enhances SR Ca<sup>2+</sup> leak that underlie asynchronous spontaneous Ca<sup>2+</sup> releases, triggering arrhythmia and impairing cardiac function.</p></div>\",\"PeriodicalId\":9678,\"journal\":{\"name\":\"Cell calcium\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.3000,\"publicationDate\":\"2024-06-17\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Cell calcium\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0143416024000836\",\"RegionNum\":2,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"CELL BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Cell calcium","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0143416024000836","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"CELL BIOLOGY","Score":null,"Total":0}
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.
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