Phosphorylation of Lamin A/C at serine 22 modulates Na_v 1.5 function

Variants in the LMNA gene, which encodes for Lamin A/C, are associated with cardiac conduction disease (CCD). We previously reported that Lamin A/C variants p.R545H and p.A287Lfs*193, which were identified in CCD patients, decreased peak I_Na in HEK-293 cells expressing Na_v 1.5. Decreased peak I_Na in the cardiac conduction system could account for patients' atrioventricular block. We found that serine 22 (Ser 22) phosphorylation of Lamin A/C was decreased in the p.R545H variant and hypothesized that lamin phosphorylation modulated Na_v 1.5 activity. To test this hypothesis, we assessed Na_v 1.5 function in HEK-293 cells co-transfected with LMNA variants or treated with the small molecule LBL1 (lamin-binding ligand 1). LBL1 decreased Ser 22 phosphorylation by 65% but did not affect Na_v 1.5 function. To test the complete loss of phosphorylation, we generated a version of LMNA with serine 22 converted to alanine 22 (S22A-LMNA); and a version of mutant R545H-LMNA that mimics phosphorylation via serine 22 to aspartic acid 22 substitution (S22D-R545H-LMNA). We found that S22A-LMNA inhibited Lamin-mediated activation of peak I_Na by 63% and shifted voltage-dependency of steady-state inactivation of Na_v 1.5. Conversely, S22D-R545H-LMNA abolished the effects of mutant R545H-LMNA on voltage-dependency but not peak I_Na . We conclude that Lamin A/C Ser 22 phosphorylation can modulate Na_v 1.5 function and contributes to the mechanism by which R545H-LMNA alters Na_v 1.5 function. The differential impact of complete versus partial loss of Ser 22 phosphorylation suggests a threshold of phosphorylation that is required for full Na_v 1.5 modulation. This is the first study to link Lamin A/C phosphorylation to Na_v 1.5 function.