Abstract
In cardiomyocytes, Na(V)1.5 channels mediate initiation and fast propagation of action potentials. The Ca(2+)-binding protein calmodulin (CaM) serves as a de facto subunit of Na(V)1.5. Genetic studies and atomic structures suggest that this interaction is pathophysiologically critical, as human mutations within the Na(V)1.5 carboxy-terminus that disrupt CaM binding are linked to distinct forms of life-threatening arrhythmias, including long QT syndrome 3, a "gain-of-function" defect, and Brugada syndrome, a "loss-of-function" phenotype. Yet, how a common disruption in CaM binding engenders divergent effects on Na(V)1.5 gating is not fully understood, though vital for elucidating arrhythmogenic mechanisms and for developing new therapies. Here, using extensive single-channel analysis, we find that the disruption of Ca(2+)-free CaM preassociation with Na(V)1.5 exerts two disparate effects: 1) a decrease in the peak open probability and 2) an increase in persistent Na(V) openings. Mechanistically, these effects arise from a CaM-dependent switch in the Na(V) inactivation mechanism. Specifically, CaM-bound channels preferentially inactivate from the open state, while those devoid of CaM exhibit enhanced closed-state inactivation. Further enriching this scheme, for certain mutant Na(V)1.5, local Ca(2+) fluctuations elicit a rapid recruitment of CaM that reverses the increase in persistent Na current, a factor that may promote beat-to-beat variability in late Na current. In all, these findings identify the elementary mechanism of CaM regulation of Na(V)1.5 and, in so doing, unravel a noncanonical role for CaM in tuning ion channel gating. Furthermore, our results furnish an in-depth molecular framework for understanding complex arrhythmogenic phenotypes of Na(V)1.5 channelopathies.