S-Nitrosylation of TRAP1 drives the progression of HFpEF through activation of the Rb-E2F axis in cardiomyocytes.

RATIONALE: Heart failure with preserved ejection fraction (HFpEF) arises from comorbidity-driven meta-inflammation and nitrosative stress, yet how these insults reprogram cardiomyocyte growth control remains unclear. We hypothesized that inducible nitric oxide synthase (iNOS)-dependent S-nitrosylation (SNO) of tumor necrosis factor receptor-associated protein 1 (TRAP1) activates E2F-driven hypertrophic programs by remodeling the retinoblastoma protein (Rb)-E2F signaling axis.

METHODS

Biotin-switch assays were used to quantify TRAP1-specific S-nitrosylation (SNO-TRAP1) in hearts from mice subjected to a two-hit HFpEF model (high-fat diet + Nω-nitro-L-arginine methyl ester (L-NAME), 12 weeks) and chow controls, as well as in cardiomyocytes, endothelial cells, and fibroblasts isolated by Langendorff perfusion. Causality was tested using pharmacological iNOS inhibition and cardiomyocyte-targeted expression of TRAP1 wild type or a S-nitrosylation cysteine 501 (Cys501) mutant. TRAP1-Rb-E2F interactions were examined using co-immunoprecipitation, domain mapping, transcriptional reporter assays, and complementary protein-protein docking analyses.

RESULTS

TRAP1 S-nitrosylation was markedly increased in HFpEF hearts, with enrichment in cardiomyocytes, and correlated with diastolic dysfunction and cardiac hypertrophy. Short-term iNOS inhibition improved diastolic indices and attenuated hypertrophic remodeling. Cardiomyocyte-specific expression of the TRAP1 Cys501A mutant reduced SNO-TRAP1 and alleviated HFpEF-like phenotypes. Mechanistically, Rb was identified as a novel TRAP1-interacting partner. SNO-TRAP1 enhanced TRAP1-Rb association while disrupting Rb-E2F binding, leading to increased E2F transcriptional activity independently of Rb phosphorylation.

CONCLUSIONS

An iNOS-SNO-TRAP1-Rb-E2F axis drives cardiomyocyte hypertrophy and diastolic dysfunction in HFpEF via phosphorylation-independent displacement of E2F from Rb. Targeting nitrosative stress or preventing TRAP1 Cys501 S-nitrosylation engagement may offer mechanism-based therapies for HFpEF.