Nav1.5 channels can reach the plasma membrane through distinct N-glycosylation states

Background: Like many voltage-gated sodium channels, the cardiac isoform Na(v)1.5 is well known as a glycoprotein which necessarily undergoes N-glycosylation processing during its transit to the plasma membrane. In some cardiac disorders, especially the Brugada syndrome (BrS), mutations in Na(v)1.5 encoding gene lead to intracellular retention and consequently trafficking defect of these proteins. We used two BrS mutants as tools to clarify both Na(v)1.5 glycosylation states and associated secretory behaviors. Methods: Patch-clamp recordings and surface biotinylation assays of HEK293T cells expressing wild-type (WT) and/or mutant Na(v)1.5 proteins were performed to assess the impact of mutant co-expression on the membrane activity and localization of WT channels. Enzymatic deglycosylation assays and brefeldin A (BEA) treatments were also employed to further characterize recombinant and native Na(v)1.5 maturation. Results: The present data demonstrate that Na(v)1.5 channels mainly exist as two differentially glycosylated forms. We reveal that dominant negative effects induced by BrS mutants upon WT channel current result from the abnormal surface expression of the fully-glycosylated forms exclusively. Furthermore, we show that coreglycosylated channels can be found at the surface membrane of BFA-treated or untreated cells, but obviously without generating any sodium current. Conclusions: Our findings provide evidence that native and recombinant Na(v)1.5 subunits are expressed as two distinct matured forms. Fully-glycosylated state of Na(v)1.5 seems to determine its functionality whereas coreglycosylated forms might be transported to the plasma membrane through an unconventional Golgiindependent secretory route. General significance: This work highlights that N-linked glycosylation processing would be critical for Na(v)1.5 membrane trafficking and function. (C) 2015 Elsevier B.V. All rights reserved.