Dissecting the mechanistic differences between Jervell Lange-Neilson syndrome type 1 and 2 using human induced pluripotent stem cell derived cardiomyocytes

Abstract Background Jervell Lange-Neilson syndrome (JLNS) is a recessive form of congenital long QT syndrome caused by mutations in either the KCNQ1 gene (JLNS-1) or the KCNE1 gene (JLNS-2). KCNQ1 encodes for the pore forming subunit of the slow delayed rectifier potassium current (IKs) and KCNE1 encodes for the regulatory subunit of IKs. Despite loss of either gene resulting in the loss of IKs, KCNQ1 mutations have a more severe clinical phenotype with higher arrhythmia burden and a longer QT prolongation compared to KCNE1 mutations. Indicating that more than just loss of IKs is responsible for the severity JLNS-1 clinical presentation. Purpose This study aimed to gain a better understanding of the differences in clinical phenotype in JLNS-1 and JLNS-2 by generating iPSC lines with either KCNQ1 or KCNE1 knock out. Methods KCNQ1 and KCNE1 knock out (KO) iPSC lines were generated using CRISPR/Cas9 technology by targeting exon 5 of KCNQ1 (the voltage sensitive domain) and Exon 4 of KCNE1 (the only protein coding exon). The iPSC KO lines were then differentiated into cardiomyocytes (CM) and action potential (AP) morphology was compared using a genetically encoded voltage sensitive reporter gene. Results KO generation was validated in both lines via DNA 0.0001). The KCNE1 KO APD (522.3 ms) was also longer than the WT (P=0.0223) but shorter than the KCNQ1 KO (P=0.0006). Interestingly the difference between KCNQ1 and KCNE1 was lost when paced at 2 Hz (P=0.3604), when IKs would be expected to be more prominent. The APD of KCNQ1 KO and KCNE1 KO remained longer than WT isogenic control at 2 Hz (Q1:P=0.0027; E1:P=0.0216). KCNQ1 KO also had a slower time to peak compared to KCNE1 KO (P=0.0036) and WT (P=0.0073) indicating altered depolarisation kinetics. At a molecular level KO of KCNE1 resulted in a 3-fold increase in expression of KCNQ1 compared to WT (P=0.0099) and KCNQ1 KO (P=0.0017) which was accompanied by increases in the expression of two genes involved in calcium handling: ATP1A2 (KCNE1 v WT: P=0.0206; KCNE1 v KCNQ1: P=0.0136) and RRAD expression (KCNE1 v WT: P=0.0428; KCNE1 v KCNQ1: P=0.0533). Conclusions Our results show key differences in AP morphology between KCNQ1 and KCNE1 KO iPSC-CM which cannot be explained by loss of IKs alone. The results also indicate a potential mechanism involving altered sodium and calcium handling genes which will be explored further.For image description, please refer to the figure legend and surrounding text.