Impact of arrhythmogenic calmodulin variants on small conductance Ca²⁺ -activated K⁺ (SK3) channels

Calmodulin (CaM) is a ubiquitous Ca²⁺ -sensing protein regulating many important cellular processes. Several CaM-associated variants have been identified in a small group of patients with cardiac arrhythmias. The mechanism remains largely unknown, even though a number of ion channels, including the ryanodine receptors and the L-type calcium channels have been shown to be functionally affected by the presence of mutant CaM. CaM is constitutively bound to the SK channel, which underlies the calcium-gated I_SK contributing to cardiac repolarization. The CaM binding to SK channels is essential for gating, correct assembly, and membrane expression. To elucidate the effect of nine different arrhythmogenic CaM variants on SK3 channel function, HEK293 cells stably expressing SK3 were transiently co-transfected with CaM^{WT or variant} and whole-cell patch-clamp recordings were performed with a calculated free Ca²⁺ concentration of 400 nmol/L. MDCK cells were transiently transfected with SK3 and/or CaM^{WT or variant} to address SK3 and CaM localization by immunocytochemistry. The LQTS-associated variants CaM^D96V , CaM^D130G , and CaM^F142L reduced I_SK,Ca compared with CaM^WT (P < 0.01, P < 0.001, and P < 0.05, respectively). The CPVT associated variant CaM^N54I also reduced the I_SK,Ca (P < 0.05), which was linked to an accumulation of SK3/CaM^N54I channel complexes in intracellular compartments (P < 0.05). The CPVT associated variants, CaM^A103V and CaM^D132E only revealed a tendency toward reduced current, while the variants CaM^F90L and CaM^N98S , causing LQTS syndrome, did not have any impact on I_SK,Ca . In conclusion, we found that the arrhythmogenic CaM variants CaM^N54I , CaM^D96V , CaM^D130G , and CaM^F142L significantly down-regulate the SK3 channel current, but with distinct mechanism.