Abstract 16449: miR-195 Regulates Myocardial SIRT3 Expression and Mitochondrial Enzyme Acetylation
Background: Unique cardiac and systemic miRNAs play an important role in cardiac remodeling and the associated response to injury by modulating key signaling elements. Through deep-sequencing of cardiac and circulating non-coding miRNAs, we identified miR-195 as the only miRNA up-regulated in plasma, serum and myocardium of patients with advanced heart failure (HF). Further, binding elements for miR-195 were found in the 3’UTR region of sirtuin3 (SIRT3), the major mitochondrial deacetylase. We hypothesized that miR-195 regulates myocardial SIRT3 expression and mitochondrial protein acetylation levels with subsequent changes in cardiac metabolism.
Methods: Cellular signaling was analyzed in human cardiomyocyte-like AC16 cells and acetylation levels in a rodent model of transgenic miR-195 overexpression were compared to WT. Luciferase assays, Western blotting and immunoprecipitation (Co-IP) assays were performed. Enzymatic activities of pyruvate dehydrogenase (PDH) and ATP synthase were measured.
Results: We observed suppression of SIRT3 and increased total protein acetylation in failing human myocardium. Luciferase assays confirmed that miR-195 directly targets the SIRT3 mRNA 3’UTR and negatively regulates SIRT3 expression. Transfection of miR-195 into AC16 cells resulted in a pronounced decrease in SIRT3 expression levels and induction of total protein acetylation levels. Myocardium of miR-195 transgenic animals was showed a global increase in total protein acetylation compared to WT. Co-IP assays revealed increased acetylation of 3 subunits of PDH (2.1-, 1.6-, 2.2-fold) and ATP synthase α subunit (2.3-fold), two key regulators of mitochondrial energy metabolism. Enhanced acetylation of these proteins correlated with a 24% decrease in PDH activity and a 30% decrease in ATP synthase activity.
Conclusions: Altogether, these data demonstrate a crucial role of miR-195 in HF and identified SIRT3 as a direct target of miR-195. Our findings suggest a new pathway of abnormal cardiac energy metabolism in the failing myocardium through miR-195-mediated SIRT3 suppression and increased protein acetylation. These changes result in specific inhibition of PDH and ATP synthase activity leading to impaired energy metabolism and ATP deprivation.
Author Disclosures: X. Zhang: None. R. Ji: None. X. Liao: None. D. Brunjes: None. E. Castillero: None. I. George: None. M. Hirano: None. P. Kennel: None. P. Schulze: None.
- © 2015 by American Heart Association, Inc.