Voltage-gated sodium channels initiate and propagate action potentials by mediating Na + influx. Nav1.4, predominantly found in the skeletal muscle, is essential for contraction and movement, and its dysfunction underlies disorders such as paramyotonia congenita and muscular dystrophy. Regulation of these channels is physiologically critical. Nav1.5, the cardiac isoform, is modulated by the small ubiquitin-like modifier protein SUMO1 at lysine 442, producing arrhythmogenic late current. Based on structural similarity, we hypothesize that Nav1.4 is also SUMOylated, with potential consequences for skeletal muscle excitability. SUMOylation is the enzymatic attachment of SUMO to lysine residues on target proteins, a modification upregulated under stress conditions such as hypoxia. To test for SUMO1 association with Nav1.4, we employed Förster Resonance Energy Transfer (FRET) microscopy in HEK293T cells. Using FRET microscopy on transfected HEK293T cells, we detected a significant interaction between Nav1.4 and SUMO1, consistent with that of Nav1.5. FRET was chosen because it directly reports molecular proximity on the nanometer scale, allowing detection of protein-protein interactions in live cells. This makes it uniquely suited for identifying SUMO1 binding events prior to electrophysiological characterization. Candidate lysine sites identified by prior mutagenesis are being tested to disrupt SUMO1 binding and refine the site of modification. These molecular interaction studies complement ongoing TEVC work to define functional consequences of SUMOylation. Together, our findings provide live-cell evidence that SUMO1 associates with Nav1.4 and highlight SUMOylation as a potential regulatory mechanism for skeletal muscle excitability.
Vishwa Madhusudhanan (Sun,) studied this question.