Abstract-Brugada syndrome is an inherited cardiac disorder caused by mutations in the cardiac sodium channel gene, SCN5A, that leads to ventricular fibrillation and sudden death. This study reports the changes in functional expression and cellular localization of an SCN5A double mutant (R1232W/T1620M) recently discovered in patients with Brugada syndrome. Mutant and wild-type (WT) human heart sodium channels (hNa v 1.5) were expressed in tsA201 cells in the presence of the ␤ 1 -auxiliary subunit. Patch-clamp experiments in whole-cell configuration were conducted to assess functional expression. Immunohistochemistry and confocal microscopy were used to determine the spatial distribution of either WT or mutant cardiac sodium channels. The results show an abolition of functional sodium channel expression of the hNa v 1.5/R1232W/T1620M mutant in the tsA201 cells. A conservative positively charged mutant, hNa v 1.5/ R1232K/T1620M, produced functional channels. Immunofluorescent staining showed that the FLAG-tagged hNa v 1.5/WT transfected into tsA201 cells was localized on the cell surface, whereas the FLAG-tagged hNa v 1.5/ R1232W/T1620M mutant was colocalized with calnexin within the endoplasmic reticulum (ER

These findings suggest that the Na(v)1.5/V1763M channel dysfunction and possible neighboring mutants contribute to a persistent inward current due to altered inactivation kinetics and clinically congenital LQTS with perinatal onset of arrhythmias that responded to lidocaine and mexiletine.

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Mammalian cells poorly express rNa v 1.8 channels.In contrast, rNa v 1.7 dorsal root ganglion channels have 90-fold higher peak Na + current densities. We investigated the role of rNa v 1.7 and rNa v 1.8 carboxy-termini in modulating the expression of rNa v 1.7 and rNa v 1.8 channels in tsA201 cells. Mutations in the ubiquitination site of the C-terminus did not improve rNa v 1.8 current levels. However, rNa v 1.8 chimeras containing the entire or the proximal portion of the rNa v 1.7 Cterminus expressed 3.2-fold and 4.8-fold higher peak current densities, respectively, than parent rNa v 1.8 channels. We conclude that the two Na + channels may have di¡erent endoplasmic reticulum processing signals. ß 2004 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.

The congenital long QT syndrome (LQTS) is a hereditary cardiac disease characterized by prolonged ventricular repolarization, syncope, and sudden death. Mutations causing LQTS have been identified in various genes that encode for ionic channels or their regulatory subunits. Several of these mutations have been reported on the KCNQ1 gene encoding for a potassium channel or its regulatory subunit (KCNE1). In this study, we report the biophysical characteristics of a new mutation (L251P) in the transmembrane segment 5 (S5) of the KCNQ1 potassium channel. Potassium currents were recorded from CHO cells transfected with either wild type or mutant KCNQ1 in the presence or in the absence of its regulatory subunit (KCNE1), using the whole-cell configuration of the patch clamp technique. Wild-type KCNQ1 current amplitudes are increased particularly by KCNE1 co-expression but no current is observed with the KCNQ1 (L251P) mutant either in the presence or in the absence of KCNE1. Coexpressing KCNE1 with equal amount of cDNAs encoding wild type and mutant KCNQ1 results in an 11-fold reduction in the amplitude of potassium currents. The kinetics of activation and inactivation and the activation curve are minimally affected by this mutation. Our results suggest that the dominant negative effect of the P251L mutation on KCNQ1 channel explains the prolonged repolarization in patients carrying this mutation.