Cardiomyocyte-Specific RNF128 Attenuates Pathological Cardiac Hypertrophy Progression by Stabilizing SERCA2a through Lys63-Linked Polyubiquitination

Abstract

Pathological cardiac hypertrophy is maladaptive cardiac remodeling induced by chronic adverse stimuli. In this study, the E3 ubiquitin ligase RNF128 was identified as a suppressor of pathological cardiac dysfunction with therapeutic value. Methods The expression of Ring Finger protein 128 (RNF128) in pathological cardiac hypertrophy was characterized via public database analysis, scRNA-seq (single-cell RNA sequencing), and further validated in clinical myocardial samples and mouse disease models. The regulatory function of RNF128 in the progression of cardiac hypertrophy was verified in vivo by cardiomyocyte-specific RNF128 knockout mice and cTnT-AAV9-mediated RNF128 overexpression. Ang II (angiotensin II)-stimulated NMCMs (neonatal mouse cardiomyocytes) were used for in vitro validation. Moreover, the downstream target of RNF128 was identified through integrated analysis of scRNA-seq, interactome profiling and quantitative proteomics, followed by a panel of molecular assays. Results Pathological cardiac hypertrophy reduced RNF128 expression in both human and murine samples. RNF128 deficiency aggravated cardiac dysfunction and pathological remodeling, while its overexpression protected cardiac function. Mechanistically, RNF128 directly interacted with SERCA2a, and catalyzed K63-linked polyubiquitination of SERCA2a at residue K476 (lysine 476), thereby impaired SERCA2a recognition by SQSTM1/p62. Consequently, RNF128 inhibited autophagy-lysosome-mediated degradation of SERCA2a. Conclusions The findings of this study highlight RNF128 as a novel therapeutic target for heart failure, linking ubiquitination-dependent protein regulation to calcium handling in cardiomyocytes.