SENP2基因敲除HEK293细胞

BACKGROUND:Decreased peak sodium current (I) and increased late sodium current (I), through the cardiac sodium channel Na1.5 encoded by SCN5A, cause arrhythmias. Many Na1.5 posttranslational modifications have been reported. A recent report concluded that acute hypoxia increases I by increasing a small ubiquitin-like modifier (SUMOylation) at K442-Na1.5. OBJECTIVE:The purpose of this study was to determine whether and by what mechanisms SUMOylation alters I, I, and cardiac electrophysiology. METHODS:SUMOylation of Na1.5 was detected by immunoprecipitation and immunoblotting. I was measured by patch clamp with/without SUMO1 overexpression in HEK293 cells expressing wild-type (WT) or K442R-Na1.5 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. Na1.5 trafficking was detected by immunofluorescence. RESULTS:Na1.5 was SUMOylated in HEK293 cells, NRCMs, and human heart tissue. HyperSUMOylation at Na1.5-K442 increased I in NRCMs and in HEK cells overexpressing WT but not K442R-Na1.5. SUMOylation did not alter other channel properties including I. AAV9-SENP2 or anacardic acid decreased I, prolonged QRS duration, and produced heart block and arrhythmias in SCN5A mice, whereas AAV9-SUMO1 increased I and shortened QRS duration. SUMO1 overexpression enhanced membrane localization of Na1.5. CONCLUSION:SUMOylation of K442-Na1.5 increases peak I without changing I, at least in part by altering membrane abundance. Our findings do not support SUMOylation as a mechanism for changes in I Na1.5 SUMOylation may modify arrhythmic risk in disease states and represents a potential target for pharmacologic manipulation.