MICU3 Regulates Mitochondrial Calcium and Cardiac Hypertrophy

Calcium (Ca ) uptake by mitochondria occurs via the mitochondrial Ca uniporter. Mitochondrial Ca uniporter exists as a complex, regulated by 3 MICU (mitochondrial Ca uptake) proteins localized in the intermembrane space: MICU1, MICU2, and MICU3. Although MICU3 is present in the heart, its role is largely unknown. We used CRISPR-Cas9 to generate a mouse with global deletion of MICU3 and an adeno-associated virus (AAV9) to overexpress MICU3 in wild-type mice. We examined the role of MICU3 in regulating mitochondrial calcium ([Ca ] ) in ex vivo hearts using an optical method following adrenergic stimulation in perfused hearts loaded with a Ca -sensitive fluorophore. Additionally, we studied how deletion and overexpression of MICU3, respectively, impact cardiac function in vivo by echocardiography and the molecular composition of the mitochondrial Ca uniporter complex via Western blot, immunoprecipitation, and Blue native-PAGE analysis. Finally, we measured MICU3 expression in failing human hearts. MICU3 knock out hearts and cardiomyocytes exhibited a significantly smaller increase in [Ca ] than wild-type hearts following acute isoproterenol infusion. In contrast, heart with overexpression of MICU3 exhibited an enhanced increase in [Ca ] compared with control hearts. Echocardiography analysis showed no significant difference in cardiac function in knock out MICU3 mice relative to wild-type mice at baseline. However, mice with overexpression of MICU3 exhibited significantly reduced ejection fraction and fractional shortening compared with control mice. We observed a significant increase in the ratio of heart weight to tibia length in hearts with overexpression of MICU3 compared with controls, consistent with hypertrophy. We also found a significant decrease in MICU3 protein and expression in failing human hearts. Our results indicate that increased and decreased expression of MICU3 enhances and reduces, respectively, the uptake of [Ca ] in the heart. We conclude that MICU3 plays an important role in regulating [Ca ] physiologically, and overexpression of MICU3 is sufficient to induce cardiac hypertrophy, making MICU3 a possible therapeutic target.