Abstract 20155: Molecular and Functional Characterization of a Novel RYR2 Mutation Linked to Long QT Syndrome
Background: Long QT (LQT) syndrome is characterized by prolongation of the QT interval in the ECG, syncope and sudden death. Mutations in 16 genes that encode ion channels or associated proteins account for ~80% of all cases, however, 20% of the cases still remain genetically unknown. Further, mutations in the cardiac ryanodine receptor gene (RYR2) have been implicated in arrhythmia syndromes such as cathecolaminergic polymorphic ventricular tachycardia (CPVT).
Results: We identified a novel mutation in RYR2 (R2920Q), in a patient with family history of sudden death, syncope and prolongation of the QT interval (QTc, 525 ms). 80 cardiogenes were simultaneously sequenced using Haloplex design on a MiSeq device. All novel or low frequency variants predicted to be damaging were confirmed by Sanger sequencing. To elucidate the mechanisms by which R2920Q might cause RyR2 channel dysfunction and lead to LQT instead of CPVT, we engineered the R2920Q mutation in the murine Ryr2 (mRyR2) and expressed it in HEK293 cells and in human iPS-derived cardiomyocytes (hIPS-CM) from a healthy control. Recombinant RyR2-R2920Q channels displayed increased Ca2+ sensitivity compared to RyR2-WT in [3H]ryanodine binding experiments (Kd for Ca2+ activation = 323 and 707 nM, respectively, n=10), with no difference in protein expression. Monolayers of purified hiPS-CM expressing mRyR2-R2920Q, mRyR2-WT or non-transfected (control) were loaded with a voltage sensitive dye and subjected to optical recordings of action potentials (AP) at 1 Hz pacing. hIPS-CM expressing mRYR2-R2920Q showed an APD90 of 267±13 compared to 197±13 and 185±29 ms in hIPS-CM expressing mRyR2-WT and non-transfected, respectively p<0.01(n=4-6). Expression of mRYR constructs in monolayers of hiPS-CM was confirmed by RT-PCR using mouse primers.
Conclusion: The novel RyR2-R2920Q mutation confers to RyR2 channels a gain of function that may be linked to the LQT syndrome in this patient. Abnormal intracellular Ca2+ handling has recently emerged as an important mechanism for arrhythmia generation in animal models of LQT. Spontaneous Ca2+ release by hyperactive RyR2 channels may give rise to LQT instead of CPVT if the release occurs during the systolic phase of the AP, a mechanism we are currently investigating.
- Cellular Electrophysiology
- Excitation-contraction coupling (ECC)
- Gene mutations
Author Disclosures: C.R. Valdivia: None. E. Antunez-Arguelles: None. J. Hernandez: None. T. Herron: None. T. Villarreal: None. P. Iturralde: None. A. Medeiros-Domingo: None. H.H. Valdivia: None.
- © 2016 by American Heart Association, Inc.