Kang Po Wei, Woodbury Lucy, Angsutararux Paweorn, Sambare Namit, Shi Jingyi, Marras Martina, Abella Carlota, Bedi Anish, Zinn DeShawn, Cui Jianmin, Silva Jonathan R
Department of Biomedical Engineering, Washington University in St.Louis, St. Louis, MO 63130, USA.
PNAS Nexus. 2023 Oct 14;2(11):pgad335. doi: 10.1093/pnasnexus/pgad335. eCollection 2023 Nov.
Missense variants in calmodulin (CaM) predispose patients to arrhythmias associated with high mortality rates ("calmodulinopathy"). As CaM regulates many key cardiac ion channels, an understanding of disease mechanism associated with CaM variant arrhythmias requires elucidating individual CaM variant effects on distinct channels. One key CaM regulatory target is the KCNQ1 (K7.1) voltage-gated potassium channel that carries the I current. Yet, relatively little is known as to how CaM variants interact with KCNQ1 or affect its function. Here, we take a multipronged approach employing a live-cell fluorescence resonance energy transfer binding assay, fluorescence trafficking assay, and functional electrophysiology to characterize >10 arrhythmia-associated CaM variants for effect on KCNQ1 CaM binding, membrane trafficking, and channel function. We identify one variant (G114W) that exhibits severely weakened binding to KCNQ1 but find that most other CaM variants interact with similar binding affinity to KCNQ1 when compared with CaM wild-type over physiological Ca ranges. We further identify several CaM variants that affect KCNQ1 and I membrane trafficking and/or baseline current activation kinetics, thereby delineating KCNQ1 dysfunction in calmodulinopathy. Lastly, we identify CaM variants with no effect on KCNQ1 function. This study provides extensive functional data that reveal how CaM variants contribute to creating a proarrhythmic substrate by causing abnormal KCNQ1 membrane trafficking and current conduction. We find that CaM variant regulation of KCNQ1 is not uniform with effects varying from benign to significant loss of function, suggesting how CaM variants predispose patients to arrhythmia via the dysregulation of multiple cardiac ion channels. : Biological, Health, and Medical Sciences, Physiology.
钙调蛋白（CaM）中的错义变异使患者易患与高死亡率相关的心律失常（“钙调蛋白病”）。由于CaM调节许多关键的心脏离子通道，因此要了解与CaM变异心律失常相关的疾病机制，就需要阐明单个CaM变异对不同通道的影响。一个关键的CaM调节靶点是携带I电流的KCNQ1（K7.1）电压门控钾通道。然而，关于CaM变异如何与KCNQ1相互作用或影响其功能，人们所知相对较少。在这里，我们采用多管齐下的方法，运用活细胞荧光共振能量转移结合试验、荧光转运试验和功能电生理学，来表征10多种与心律失常相关的CaM变异对KCNQ1的CaM结合、膜转运和通道功能的影响。我们鉴定出一种变异体（G114W），它与KCNQ1的结合严重减弱，但发现与生理钙范围内的野生型CaM相比，大多数其他CaM变异与KCNQ1的结合亲和力相似。我们进一步鉴定出几种影响KCNQ1和I膜转运和/或基线电流激活动力学的CaM变异体，从而阐明钙调蛋白病中KCNQ1的功能障碍。最后，我们鉴定出对KCNQ1功能无影响的CaM变异体。这项研究提供了大量的功能数据，揭示了CaM变异如何通过导致KCNQ1膜转运异常和电流传导异常，从而促成心律失常底物的形成。我们发现CaM变异对KCNQ1的调节并不一致，其影响从良性到功能严重丧失不等，这表明CaM变异如何通过多种心脏离子通道的失调使患者易患心律失常。 ：生物、健康和医学科学，生理学。
