Abstract Body: Cardiovascular disease remains the leading cause of death worldwide, accounting for nearly 18 million deaths annually. Myocardial injury causes irreversible cardiomyocyte loss and triggers inflammatory and reparative responses that replace functional myocardium with fibrotic scar tissue, culminating in cardiac hypertrophy, fibrosis, ventricular stiffening, dilation, and ultimately heart failure. Despite its clinical significance, effective strategies to prevent adverse cardiac remodeling or halt HF progression remain lacking. Recent advances in epitranscriptomics highlight the importance of RNA modifications in regulating gene expression and stress responses. Pseudouridylation, the most abundant RNA modification, influences RNA stability, translation, and RNA-protein interactions. Among pseudouridine synthases (Pus), Pus7 stands out as one of the principal mRNA-modifying enzymes. We recently demonstrated that Pus7 modulates the pseudouridylation of histone mRNAs and regulates RNA autophagy in kidney cells, whereas its functional role in cardiovascular diseases remains unknown. Here, we define the roles of PUS7 in pathological cardiac remodeling using complementary cellular and murine models. In cultured cardiomyocytes, Pus7 knockdown significantly attenuated phenylephrine-induced pathological cardiac hypertrophy, as demonstrated by reduced cell size and decreased hypertrophic gene expression. In cardiac fibroblasts, Pus7 silencing markedly suppressed fibrotic gene expression and mitigated TGFβ-induced fibroblast activation. In vivo, transgenic Pus7-deficient mice exhibited significantly attenuated adverse cardiac remodeling and improved cardiac functions in both myocardial infarction and pressure overload models. Mechanistically, Pus7 silencing robustly activated autophagy flux in both cardiomyocytes and cardiac fibroblasts, as evidenced by elevated LC3B-II levels in the presence of bafilomycin A and increased autophagosome maturation assessed by the autophagic flux reporter assays. Collectively, these findings identify PUS7 as a previously unrecognized regulator of adverse cardiac remodeling. Targeting PUS7-mediated pseudouridylation may represent a novel therapeutic strategy to enhance autophagy, mitigate cardiac fibrosis and hypertrophy, and ultimately prevent heart failure progression.