Divergent Regulation of RyR2 Calcium Release Channels by Arrhythmogenic Human Calmodulin Missense Mutants
Rationale: Calmodulin (CaM) mutations are associated with an autosomal-dominant syndrome of ventricular arrhythmia and sudden death that can present with divergent clinical features of catecholaminergic polymorphic ventricular tachycardia (CPVT) or long QT syndrome (LQTS). CaM binds to and inhibits RyR2 Ca release channels in the heart, but whether arrhythmogenic CaM mutants alter RyR2 function is not known.
Objective: To gain mechanistic insight into how human CaM mutations affect RyR2 Ca channels.
Methods and Results: We studied recombinant CaM mutants associated with CPVT (N54I, N98S) or LQTS (D96V, D130G, F142L). As a group, all LQTS-associated CaM mutants (LQTS-CaMs) exhibited reduced Ca affinity, whereas CPVT-associated CaM mutants (CPVT-CaMs) had either normal or modestly lower Ca affinity. In permeabilized ventricular myocytes, CPVT-CaMs at a physiological intracellular concentration (100nM) promoted significantly higher spontaneous Ca wave and spark activity, a typical cellular phenotype of CPVT. Compared to wild-type (WT) CaM, CPVT-CaMs caused greater RyR2 single channel open probability and showed enhanced binding affinity to RyR2. Even a 1:8 mixture of CPVT-CaM:WT-CaM activated Ca waves, demonstrating functional dominance. By contrast, LQTS-CaMs did not promote Ca waves and exhibited either normal regulation of RyR2 single channels (D96V) or lower RyR2 binding affinity (D130G, F142L). None of the CaM mutants altered Ca/CaM binding to CaM-kinase II.
Conclusions: A small proportion of CPVT-CaM is sufficient to evoke arrhythmogenic Ca disturbances, whereas LQTS-CaMs do not. Our findings explain the clinical presentation and autosomal dominant inheritance of CPVT-CaM mutations and suggest that RyR2-interactions are unlikely to explain arrhythmogenicity of LQTS-CaM mutations.
- RyR2 Calcium Release Channel
- Catecholaminargic Polymorphic Ventricular Tachycardia
- long QT syndrome
- sarcoplasmic reticulum
- ryanodine receptor
- calcium sparks
- calcium/calmodulin-dependent protein kinase II
- Received January 6, 2014.
- Revision received February 19, 2014.
- Accepted February 21, 2014.