Structural basis of human Na_v1.5 gating mechanisms

Voltage-gated Na_v1.5 channels are central to the generation and propagation of cardiac action potentials¹. Aberrations in their function are associated with a wide spectrum of cardiac diseases including arrhythmias and heart failure^2-5. Despite decades of progress in Na_v1.5 biology^6-8, the lack of structural insights into intracellular regions has hampered our understanding of its gating mechanisms. Here we present three cryo-EM structures of human Na_v1.5 in previously unanticipated open states, revealing sequential conformational changes in gating charges of the voltage-sensing domains (VSDs) and several intracellular regions. Despite the channel being in the open state, these structures show the IFM motif repositioned in the receptor site but not dislodged. In particular, our structural findings highlight a dynamic C-terminal domain (CTD) and III-IV linker interaction, which regulates the conformation of VSDs and pore opening. Electrophysiological studies confirm that disrupting this interaction results in the fast inactivation of Na_v1.5. Together, our structure-function studies establish a foundation for understanding the gating mechanisms of Na_v1.5 and the mechanisms underlying CTD-related channelopathies.