Long QT Syndrome KCNH2 Variant Induces hERG1a/1b Subunit Imbalance in Patient-Specific Induced Pluripotent Stem Cell-Derived Cardiomyocytes.
Long QT Syndrome KCNH2 Variant Induces hERG1a/1b Subunit Imbalance in Patient-Specific Induced Pluripotent Stem Cell-Derived Cardiomyocytes.
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DOI：
10.1161/circep.120.009343
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发表时间：
2021-04
期刊：
影响因子：
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通讯作者：
Kamp TJ
中科院分区：
文献类型：
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作者：
Feng L;Zhang J;Lee C;Kim G;Liu F;Petersen AJ;Lim E;Anderson CL;Orland KM;Robertson GA;Eckhardt LL;January CT;Kamp TJ
遗传性长QT综合征2型(LQT2)是由编码hERG1钾通道的KCNH2基因变异引起的。两种主要的异构体hERG1a和hERG1b聚集形成四聚体通道。N-末端Per-Arnt-Sim(PAS)结构域只存在于hERG1a亚基上，是致病变异体的研究热点，但PAS结构域变异体是否影响hERG1b的表达与LQT2表型有关尚不清楚。我们的目的是利用患者特异性诱导多能干细胞来源的心肌细胞(IPSC-CMS)来研究hERG1a PAS结构域变异体hERG1-H70R的发病机制。人IPSCs来自一名携带PAS结构域变体hERG1-H70R的LQT2患者。CRISPR/Cas9基因编辑产生了等基因对照IPSC系。对分化的IPSC-CMS进行电生理、hERG1a/1b基因表达和hERG1a/1b蛋白表达的检测。与对照组相比，单个hERG1-H70R IPSC-CMS的动作电位延长，电压钳研究显示IKR的潜在降低与加速失活有关。在hERG1-H70R IPSC-CMS中，hERG1a和hERG1b的转录水平与对照组相比没有明显变化，但hERG1b的转录水平和hERG1b/hERG1a的比例显著增加，提示转录后的变化。HERG1-H70R IPSC-CMS中复杂糖基化hERG1a的表达因蛋白质转运受损而降低，而复杂糖基化形式的hERG1b的表达没有变化。患者特异性的hERG1-H70R IPSC-CMS揭示了一种新的LQT2表型的发病机制，这是由于hERG1a的运输受损和hERG1b的表达维持，从而产生亚单位失衡，并通过加速失活减少IKR。
Inherited long QT syndrome type 2 (LQT2) results from variants in the KCNH2 gene encoding the hERG1 potassium channel. Two main isoforms, hERG1a and hERG1b, assemble to form tetrameric channel. The N-terminal Per-Arnt-Sim (PAS) domain, present only on hERG1a subunits, is a hotspot for pathogenic variants, but it is unknown whether PAS domain variants impact hERG1b expression to contribute to the LQT2 phenotype. We aimed to use patient-specific induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to investigate the pathogenesis of the hERG1a PAS domain variant hERG1-H70R. Human iPSCs were derived from a LQT2 patient carrying the PAS domain variant hERG1-H70R. CRISPR/Cas9 gene editing produced isogenic control iPSC lines. Differentiated iPSC-CMs were evaluated for their electrophysiology, hERG1a/1b mRNA expression, and hERG1a/1b protein expression. Action potentials from single hERG1-H70R iPSC-CMs were prolonged relative to controls, and voltage clamp studies showed an underlying decrease in IKr with accelerated deactivation. In hERG1-H70R iPSC-CMs, transcription of hERG1a and hERG1b mRNA was unchanged compared to controls based on nascent nuclear transcript analysis, but hERG1b mRNA was significantly increased as was the ratio of hERG1b/hERG1a in mRNA complexes, suggesting post-transcriptional changes. Expression of complex glycosylated hERG1a in hERG1-H70R iPSC-CMs was reduced due to impaired protein trafficking, whereas the expression of the complex glycosylated form of hERG1b was unchanged. Patient-specific hERG1-H70R iPSC-CMs reveal a newly appreciated mechanism of pathogenesis of the LQT2 phenotype due to both impaired trafficking of hERG1a and maintained expression of hERG1b that produces subunit imbalance and reduced IKr with accelerated deactivation.