RND3 Enhances Cardiac Glucose Metabolism Through Inhibiting ACAT1-Dependent PDHA1 Acetylation and Protects Against Ischemia-Reperfusion Injury.

Abstract

Metabolic disturbances are key contributors to myocardial ischemia-reperfusion (I/R) injury, yet the underlying molecular mechanisms remain largely unclear. RND3, a cytosolic small GTPase known to antagonize ROCK1 (Rho-associated coiled-coil kinase 1), has been implicated in several cardiovascular disorders. However, its mitochondrial localization and functional role in cardiac energy metabolism and I/R injury remain unknown. A murine model of myocardial I/R injury was established through left anterior descending coronary artery ligation. Mice with cardiomyocyte-specific knockout and overexpression of <i>Rnd3</i> were generated. To investigate the role of RND3 in cardiac metabolism and I/R injury, we used ¹³C-nuclear magnetic resonance, ¹⁸F-fluorodeoxyglucose positron emission tomography/computed tomography scanning, seahorse mitochondrial energy metabolism assays, and ¹³C-metabolic flux tracing. Mechanistic studies were conducted using RNA sequencing, coimmunoprecipitation, mass spectrometry, and GST pulldown assays. Cardiomyocyte-specific deletion of <i>Rnd3</i> (<i>Rnd3</i>^cKO) resulted in impaired glucose oxidation and compensatory upregulation of fatty acid oxidation, leading to pronounced cardiac dysfunction and increased mortality. <i>Rnd3</i>^cKO hearts exhibited reduced pyruvate/malate-driven complex I respiration and marked uncoupling between glycolysis and the tricarboxylic acid cycle. Mechanistically, RND3 was identified as a novel mitochondrial matrix-localized small GTPase that directly binds to ACAT1 (acetyl-CoA acetyltransferase), disrupting its interaction with PDHA1 (pyruvate dehydrogenase E1α subunit) and thereby promoting PDHA1 acetylation and glucose oxidation. It is important to note that RND3 expression was significantly downregulated in both human and murine hearts after I/R insult. Loss of RND3 sensitized the hearts to I/R injury, as evidenced by reduced levels of phosphocreatine and ATP. Conversely, cardiac-specific overexpression of <i>Rnd3</i> conferred protection against I/R injury, an effect that was abolished upon <i>Pdha1</i> knockdown. Our results identify RND3 as a novel mitochondria-localized regulator of glucose oxidation that safeguards the heart against I/R injury. Therapeutic reconstitution of <i>Rnd3</i> may represent a promising strategy to restore metabolic homeostasis and mitigate myocardial damage in the context of I/R.