Brugada syndrome (BrS) is an autosomal-inherited cardiac arrhythmia characterized by an ST-segment elevation in the right precordial leads of the electrocardiogram and an increased risk of syncope and sudden death. SCN5A, encoding the cardiac sodium channel Na_v1.5, is the main gene involved in BrS. Despite the fact that several mutations have been reported in the N-terminus of Na_v1.5, the functional role of this region remains unknown. We aimed to characterize two BrS N-terminal mutations, R104W and R121W, a construct where this region was deleted, ΔNter, and a construct where only this region was present, Nter.

Patch-clamp recordings in HEK293 cells demonstrated that R104W, R121W, and ΔNter abolished the sodium current I_Na. Moreover, R104W and R121W mutations exerted a strong dominant-negative effect on wild-type (WT) channels. Immunocytochemistry of rat neonatal cardiomyocytes revealed that both mutants were mostly retained in the endoplasmic reticulum and that their co-expression with WT channels led to WT channel retention. Furthermore, co-immunoprecipitation experiments showed that Na_v1.5-subunits were interacting with each other, even when mutated, deciphering the mutation dominant-negative effect. Both mutants were mostly degraded by the ubiquitin–proteasome system, while ΔNter was addressed to the membrane, and Nter expression induced a two-fold increase in I_Na. In addition, the co-expression of N-terminal mutants with the gating-defective but trafficking-competent R878C-Na_v1.5 mutant gave rise to a small I_Na.

This study reports for the first time the critical role of the Na_v1.5 N-terminal region in channel function and the dominant-negative effect of trafficking-defective channels occurring through α-subunit interaction.