Characterization of a novel SCN5A mutation associated with Brugada syndrome reveals involvement of DIIIS4–S5 linker in slow inactivation

OBJECTIVE: Mutations in SCN5A, the gene encoding the alpha-subunit of the cardiac sodium channel (Na(v)1.5), have been associated with various inherited arrhythmia syndromes, including Brugada syndrome (BrS). Here, we report the functional consequences of a novel missense SCN5A mutation, G1319V, identified in a BrS patient. The G1319V mutation is located in the loop connecting transmembrane segments 4 and 5 in domain III (DIIIS4-S5), a region so far considered to be exclusively involved in fast inactivation. METHODS: Whole-cell mutant (G1319V) and wild-type (WT) sodium currents (I(Na)) were studied in the Human Embryonic Kidney cell line (HEK-293) transfected with Na(v)1.5 alpha-subunit cDNA (WT or mutant) together with h beta(1)-subunit cDNA, using the patch-clamp technique. RESULTS: Maximal peak I(Na) and persistent sodium current were similar in WT and channel G1319V channels. The G1319V mutation shifted the potential of half-maximal (V(1/2)) activation towards more positive potentials (+3.7 mV), thereby increasing the degree of depolarization required for activation. The V(1/2) of inactivation of G1319V channels was shifted by -6.0 mV compared to WT, resulting in a reduced channel availability. The change in the steady-state inactivation was completely due to a negative shift (-6.8 mV) of the voltage-dependence of slow inactivation, while the voltage-dependence of fast inactivation was unaffected. The fast component of recovery from inactivation of G1319V channels was slowed down. Finally, the G1319V mutation caused a two-fold increase in the propensity of the channels to enter the slow inactivated state. Reduction in I(Na) peak amplitude on repetitive depolarizations at short interpulse intervals (40 ms) was significantly more pronounced in G1319V compared to WT. Accordingly, carriers of the G1319V mutation showed marked QRS widening upon increases in heart rate during exercise testing, pointing to enhancement of slow inactivation. CONCLUSIONS: We identified the DIIIS4-S5 linker as a new region involved in slow inactivation of Na(v)1.5. The biophysical alterations of the G1319V mutation all contribute to a reduction in I(Na), in line with the proposed mechanism underlying BrS.