Abstract Fri081: SIRT5 Dysfunction Evokes Arrhythmias and Na ⁺ and Ca ²⁺ Mishandling in mouse heart

Background: The cardiac Na + channel Na V 1.5 (encoded by SCN5A ) governs inward Na + current ( I Na ), playing a crucial role in the fast upstroke and plateau phases of the cardiac action potential. Mutations in SCN5A are implicated in inherited and acquired arrhythmias, including ~20% of Brugada Syndrome (BrS) cases. Altered I Na impact Ca 2+ handling and excitation-contraction coupling. Previously, we demonstrated that SIRT1-mediated deacetylation of Na V 1.5 at K1479 enhances peak I Na . Recently, potential mutations in SIRT5—including P114T—were identified in small families with BrS. SIRT5, another NAD + -dependent deACYLase from the Sirtuin family, is primarily localized to mitochondria. Hypothesis: Sirt5 dysfunction leads to arrhythmogenesis through Na + and Ca 2+ mishandling in an oxidative stress- dependent manner in mouse hearts. Aims: To investigate SIRT5’s role in BrS using heterologous expression systems, homozygous P114T-Sirt5 knock-in (P114T-KI) and Sirt5 -/- mice. Methods: Protein expression and physical interactions were assessed by immunoprecipitation and immunoblot. Patch-clamp electrophysiology in HEK cells and mouse ventricular cardiac myocytes evaluated I Na . Reactive oxygen species (ROS) levels and Ca 2+ imaging were measured by confocal microscopy. Results: Both WT and P114T-SIRT5 co-immunoprecipitate with Na V 1.5, but only WT enhanced peak I Na in K1479-Na V 1.5 dependent manner in HEK cells. P114T-SIRT5 failed to do so. The late I Na of cardiomyocytes was elevated ( Figure, A ), and QTc was prolonged in both Sirt5 -/- and P114T mice. P114T-KI hearts showed increased Na + /Ca 2+ exchanger (NCX1) expression, elevated phosphorylation of phospholamban at Thr17 (p-PLN-Thr17, Figure, B ). Live-cell staining using DCFDA or mitoSOX revealed that ROS levels were higher in P114T-KI hearts at baseline compared to WT littermates and were further exacerbated by H 2 O 2 . Aberrant Ca 2+ handling and arrhythmias in Langendorff-perfused hearts of P114T mice were mitigated by the mitochondrial ROS scavenger MitoTEMPO ( Figure, C ). Conclusion: SIRT5 dysfunction in P114T-SIRT5 disrupts Na + and Ca 2+ homeostasis and induces arrhythmias in a ROS-dependent manner, suggesting potential mechanisms underlying BrS. These findings highlight SIRT5 as a promising therapeutic target for BrS.