RCAN1 Suppresses Mitochondrial Fission, Protecting the Heart from Calpain‐mediated Damage

Abstract

Altered calcium handling and mitochondrial function are hallmarks of heart disease and both influence heart susceptibility to ischemia reperfusion (I/R). Regulator of Calcineurin 1 (RCAN1) is a feedback inhibitor of calcineurin, a calcium-activated phosphatase involved in cardiac remodeling. We used advanced imaging techniques and an in vitro model of I/R (sim-I/R) to dissect RCAN1's role in mitochondrial dynamics and cell survival. SiRNA was used to deplete RCAN1 from neonatal rat ventricular myocytes (NRVMs). Confocal imaging showed an increase in the number of mitochondria per cell and a decreased individual mitochondrial size, indicative of increased fission in the absence of RCAN1 (See Figure). There was a corresponding decrease in the phosphorylation of the fission protein DRP1, a calcineurin substrate. Consistent with increased fission, the siRCAN1-depleted NRVMs showed decreases in ATP production, mitochondrial membrane potential, oxygen consumption, and mitochondrial calcium uptake. Death following sim-I/R was significantly greater in siRCAN1-depleted NRVMs compared to controls. Importantly, we found an increase in both cytosolic calcium levels and calpain activity in the SiRCAN1-depleted NRVMs subjected to sim-I/R. Resistance to I/R was restored in siRCAN1-depleted NRVMs by pharmacological inhibition of calcineurin (FK506), mitochondrial fission (Mdivi), or calpain. We hypothesize that RCAN1 preserves mitochondrial fusion by inhibiting calcineurin-dependent translocation of DRP1 to mitochondria, preserving mitochondrial network connectivity and cacium buffering capacity. These findings have important implications regarding cell metabolism and survival in all tissues. Funding: AHA and NIH