BACKGROUND The KCNQ1+KCNE1 (IKs) potassium channel plays a crucial role in cardiac adaptation to stress, in which β-adrenergic stimulation phosphorylates the IKs channel through the cyclic adenosine monophosphate (cAMP)/PKA (protein kinase A) pathway. Phosphorylation increases the channel current and accelerates repolarization to adapt to an increased heart rate. Variants in KCNQ1 can cause long-QT syndrome type 1 (LQT1), and those with defective cAMP effects predispose patients to the highest risk of cardiac arrest and sudden death. However, the molecular connection between IKs channel phosphorylation and channel function, as well as why high-risk LQT1 mutations lose cAMP sensitivity, remain unclear. METHODS Regular patch clamp and voltage clamp fluorometry techniques were utilized to record pore opening and voltage sensor movement of wild-type and mutant KCNQ1/IKs channels. The clinical phenotypic penetrance of each LQT1 mutation was analyzed as a metric for assessing their clinical risk. The patient-specific-induced pluripotent stem-cell model was used to test mechanistic findings in physiological conditions. RESULTS By systematically elucidating mechanisms of a series of LQT1 variants that lack cAMP sensitivity, we identified molecular determinants of IKs channel regulation by phosphorylation. These key residues are distributed across the N-terminus of KCNQ1 extending to the central pore region of IKs. We refer to this pattern as the IKs channel PKA phosphorylation axis. Next, by examining LQT1 variants from clinical databases containing 10 579 LQT1 carriers, we found that the distribution of the most high-penetrance LQT1 variants extends across the IKs channel PKA phosphorylation axis, demonstrating its clinical relevance. Furthermore, we found that a small molecule, ML277, which binds at the center of the phosphorylation axis, rescues the defective cAMP effects of multiple high-risk LQT1 variants. This finding was then tested in high-risk patient-specific induced pluripotent stem cell-derived cardiomyocytes, where ML277 remarkably alleviates the beating abnormalities. CONCLUSIONS Our findings not only elucidate the molecular mechanism of PKA-dependent IKs channel phosphorylation but also provide an effective antiarrhythmic strategy for patients with high-risk LQT1 variants.

中文翻译：

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靶向 IKs 通道 PKA 磷酸化轴以恢复其在高风险 LQT1 变异体中的功能。


背景 KCNQ1+KCNE1 (IKs) 钾通道在心脏应激适应中发挥着至关重要的作用，其中 β-肾上腺素能刺激通过环磷酸腺苷 (cAMP)/PKA（蛋白激酶 A）途径磷酸化 IKs 通道。磷酸化增加通道电流并加速复极化以适应增加的心率。 KCNQ1 的变异可导致 1 型长 QT 综合征 (LQT1)，而 cAMP 效应缺陷的变异会使患者面临最高的心脏骤停和猝死风险。然而，IKs 通道磷酸化和通道功能之间的分子联系，以及为什么高风险 LQT1 突变会失去 cAMP 敏感性，仍不清楚。方法利用常规膜片钳和电压钳荧光测定技术来记录野生型和突变型KCNQ1/IKs通道的孔开放和电压传感器运动。分析每个 LQT1 突变的临床表型外显率，作为评估其临床风险的指标。患者特异性诱导多能干细胞模型用于测试生理条件下的机制发现。结果通过系统地阐明一系列缺乏 cAMP 敏感性的 LQT1 变体的机制，我们确定了通过磷酸化调节 IK 通道的分子决定因素。这些关键残基分布在 KCNQ1 的 N 末端，延伸至 IK 的中心孔区域。我们将此模式称为 IKs 通道 PKA 磷酸化轴。接下来，通过检查包含 10 579 个 LQT1 携带者的临床数据库中的 LQT1 变体，我们发现外显率最高的 LQT1 变体的分布延伸到 IKs 通道 PKA 磷酸化轴，证明了其临床相关性。 此外，我们发现一种小分子 ML277 结合在磷酸化轴的中心，可以挽救多种高风险 LQT1 变体的缺陷 cAMP 效应。随后在高危患者特异性诱导多能干细胞衍生的心肌细胞中测试了这一发现，其中 ML277 显着减轻了跳动异常。结论 我们的研究结果不仅阐明了 PKA 依赖性 IK 通道磷酸化的分子机制，而且为高危 LQT1 变异患者提供了有效的抗心律失常策略。