Abstract
BACKGROUND: Decreased peak sodium current (I(Na)) and increased late sodium current (I(Na,L)), through the cardiac sodium channel Na(V)1.5 encoded by SCN5A, cause arrhythmias. Many Na(V)1.5 posttranslational modifications have been reported. A recent report concluded that acute hypoxia increases I(Na,L) by increasing a small ubiquitin-like modifier (SUMOylation) at K442-Na(V)1.5. OBJECTIVE: The purpose of this study was to determine whether and by what mechanisms SUMOylation alters I(Na), I(Na,L), and cardiac electrophysiology. METHODS: SUMOylation of Na(V)1.5 was detected by immunoprecipitation and immunoblotting. I(Na) was measured by patch clamp with/without SUMO1 overexpression in HEK293 cells expressing wild-type (WT) or K442R-Na(V)1.5 and in neonatal rat cardiac myocytes (NRCMs). SUMOylation effects were studied in vivo by electrocardiograms and ambulatory telemetry using Scn5a heterozygous knockout (SCN5A(+/-)) mice and the de-SUMOylating protein SENP2 (AAV9-SENP2), AAV9-SUMO1, or the SUMOylation inhibitor anacardic acid. Na(V)1.5 trafficking was detected by immunofluorescence. RESULTS: Na(V)1.5 was SUMOylated in HEK293 cells, NRCMs, and human heart tissue. HyperSUMOylation at Na(V)1.5-K442 increased I(Na) in NRCMs and in HEK cells overexpressing WT but not K442R-Na(v)1.5. SUMOylation did not alter other channel properties including I(Na,L). AAV9-SENP2 or anacardic acid decreased I(Na), prolonged QRS duration, and produced heart block and arrhythmias in SCN5A(+/-) mice, whereas AAV9-SUMO1 increased I(Na) and shortened QRS duration. SUMO1 overexpression enhanced membrane localization of Na(V)1.5. CONCLUSION: SUMOylation of K442-Na(v)1.5 increases peak I(Na) without changing I(Na,L), at least in part by altering membrane abundance. Our findings do not support SUMOylation as a mechanism for changes in I(Na,L.) Na(v)1.5 SUMOylation may modify arrhythmic risk in disease states and represents a potential target for pharmacologic manipulation.