Abstract 14602: Physical and Biophysical Coupling of the Cardiac Sodium Channel Involves 14-3-3
Introduction: Mutations in SCN5A, the gene encoding for the cardiac sodium channel, produce alterations of the cardiac action potential that lead to life-threatening arrhythmias such as Long QT Syndrome (LQT3) and Brugada Syndrome (BrS). The conventional wisdom that sodium channels exist in complexes containing a single alpha-subunit has been challenged by the existence of dominant-negative (DN) mutations in BrS and the presence of polymorphisms that can restore trafficking and gating deficiencies of mutant channels in LQT and BrS. In fact, we have previously demonstrated that SCN5A subunits can interact with each other. Here we hypothesized that the physical and biophysical interactions between SCN5A alpha-subunits involve the partner protein 14-3-3, known to form dimers.
Methods: SCN5A DN-BrS mutants and LQT3 gating deficient mutants were expressed in HEK293 cells and in commercially available iPS-derived cardiomyocytes, iCells©, in presence or absence of 14-3-3 inhibition. Resulting currents were measured using patch-clamp.
Results: In order to investigate if the DN-effect seen by some BrS mutants is due to interaction of the sodium channel with the protein 14-3-3 which in turn would be involved in the alpha-alpha interaction, we expressed two different BrS DN-mutants in HEK293 cells with and without difopein, a specific 14-3-3 inhibitor. The presence of difopein abolished the DN-effect of both mutants. The DN-effect was also abolished when we mutated the putative 14-3-3 binding site on SCN5A and expressed the DN-mutants either in HEK293 cells or in iCells©. Inhibition of 14-3-3 also impaired the biophysical coupling observed in presence of SCN5A gating deficient mutants that affect either activation or inactivation of not only the mutants but also of the wild-type channel. Conclusions: Our results suggest that binding of 14-3-3 to the cardiac sodium channel alpha-subunit is involved in the alpha-alpha interaction and biophysical coupling of the channel. This study not only shifts paradigms in regards to sodium channel assembly and structure, but also puts forward the idea that physical and biophysical uncoupling of cardiac sodium channels could be a new therapy target for cardiac arrhythmias caused by SCN5A mutations.
Author Disclosures: J.C. Clatot: None. M. Hoshi: None. H. Liu: None. X. Wan: None. K. Shinlapawittayatorn: None. E. Ficker: None. I. Deschênes: None.
- © 2015 by American Heart Association, Inc.