Rationale:
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a rare disease, manifested by syncope or sudden death in children or young adults under stress conditions. Mutations in the Ca
2+
release channel/ryanodine receptor (RyR2) gene account for about 60% of the identified mutations. Recently, we found and described a mutation in RyR2 N-terminal domain, RyR2
R420Q
.
Objective:
To determine the arrhythmogenic mechanisms of this mutation.
Methods and Results:
Ventricular tachycardias under stress conditions were observed in both CPVT patients and KI mice. During action potential recording (by patch-clamp in KI mouse cardiomyocytes and by microelectrodes in mutant hiPSC-CM) we observed an increased occurrence of delayed after-depolarizations (DADs) under isoproterenol stimulation, associated with increased Ca
2+
waves during confocal Ca
2+
recording in both mouse and human RyR2
R420Q
cardiomyocytes. In addition, Ca
2+
-induced Ca
2+
-release, as well as a rough indicator of fractional Ca
2+
release, were higher and Ca
2+
sparks longer in the RyR2
R420Q
expressing cells. At the ultrastructural nanodomain level, we observed smaller RyR2 clusters and widened junctional sarcoplasmic reticulum (jSR) measured by g-STED super-resolution and electronic microscopy, respectively. The increase in jSR width might be due to the impairment of RyR2
R420Q
binding to junctophilin-2, as there were less junctophilin-2 co-immunoprecipitated with RyR2
R420Q
. At the single current level, the RyR2R420Q channel dwells longer in the open state at low [Ca
2+
]
i
, but there is predominance of a subconductance state. The latter might be correlated with an enhanced interaction between the N-terminus and the core solenoid, a RyR2 inter-domain association that has not been previously implicated in the pathogenesis of arrhythmias and sudden cardiac death.
Conclusions:
The RyR2
R420Q
CPVT mutation modifies the interdomain interaction of the channel and weaken its association with junctophillin-2. These defects may underlie both nanoscale disarrangement of the dyad and channel dysfunction.