Cardiomyocyte‐Enriched USP20 Ameliorates Pathological Cardiac Hypertrophy by Targeting STAT3 Deubiquitination

Abstract Although pathological cardiac hypertrophy is a key driver of heart failure, the underlying mechanisms remain incompletely elucidated. This study investigates the role and mechanism of deubiquitinating enzyme (DUB) ubiquitin‐specific protease 20 (USP20) in cardiac hypertrophy. Transcriptomic profiling of hypertrophic hearts shows significant alterations in the expression of DUBs, including a remarkable downregulation of USP20. USP20 is predominantly expressed in cardiomyocytes. Co‐immunoprecipitation (Co‐IP) followed by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) is used to identify USP20 substrates. Cleavage Under Targets and Tagmentation assay (CUT&Tag) sequencing is employed to identify downstream targets of signal transducer and activator of transcription 3 (STAT3). Functionally, USP20 deficiency exacerbates cardiac hypertrophy induced by either angiotensin II (Ang II) or transverse aortic constriction (TAC), whereas USP20 overexpression alleviates hypertrophic responses. Mechanistically, USP20 deubiquitinates STAT3 by removing K63‐linked ubiquitin chains at K177 via its H645 active site, reducing STAT3 phosphorylation and nuclear translocation. This inhibites STAT3's transcriptional activity at coactivator‐associated arginine methyltransfer ( Carm1) promoter, leading to upregulated CARM1 expression and mitigated hypertrophy. Importantly, the STAT3 inhibitor Stattic confirms STAT3 serves as a key substrate mediating the cardiac protective effects of USP20. These findings unveil a novel USP20/STAT3/CARM1 axis in cardiomyocytes and reveal its therapeutic potential for cardiac hypertrophy.