Calmodulin mutations affecting Gly114 impair binding to the Na_V1.5 IQ-domain

Missense variants in CALM genes encoding the Ca²⁺-binding protein calmodulin (CaM) cause severe cardiac arrhythmias. The disease mechanisms have been attributed to dysregulation of RyR2, for Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT) and/or Ca_V1.2, for Long-QT Syndrome (LQTS). Recently, a novel CALM2 variant, G114R, was identified in a mother and two of her four children, all of whom died suddenly while asleep at a young age. The G114R variant impairs closure of Ca_V1.2 and RyR2, consistent with a CPVT and/or mild LQTS phenotype. However, the children carrying the CALM2 G114R variant displayed a phenotype commonly observed with variants in Na_V1.5, i.e., Brugada Syndrome (BrS) or LQT3, where death while asleep is a common feature. We therefore hypothesized that the G114R variant specifically would interfere with Na_V1.5 binding. Here, we demonstrate that CaM binding to the Na_V1.5 IQ-domain is severely impaired for two CaM variants G114R and G114W. The impact was most severe at low and intermediate Ca²⁺ concentrations (up to 4 µM) resulting in more than a 50-fold reduction in Na_V1.5 binding affinity, and a smaller 1.5 to 11-fold reduction at high Ca²⁺ concentrations (25-400 µM). In contrast, the arrhythmogenic CaM-N98S variant only induced a 1.5-fold reduction in Na_V1.5 binding and only at 4 µM Ca²⁺. A non-arrhythmogenic I10T variant in CaM did not impair Na_V1.5 IQ binding. These data suggest that the interaction between Na_V1.5 and CaM is decreased with certain CaM variants, which may alter the cardiac sodium current, I_Na. Overall, these results suggest that the phenotypic spectrum of calmodulinopathies may likely expand to include BrS- and/or LQT3-like traits.