RhoA-Mitophagy raw WB images

This research investigated the role of RhoA, a small G-protein, in cardioprotection against ischemic stress and showed that RhoA regulates the PINK1 stability to stimulate mitophagy, providing a cell survival advantage.Mitophagy removes dysfunctional mitochondria and is an essential process contributing to mitochondrial quality control. We tested the hypothesis that signaling through RhoA, a small G-protein, induces mitophagy via modulation of the PINK1/Parkin pathway to provide cardioprotection against ischemic stress. We demonstrate that activation of RhoA in cardiomyocytes results in an accumulation of PINK1 at mitochondria. This is accompanied by translocation of Parkin to mitochondria and ubiquitination of mitochondrial proteins, leading to elimination of damaged mitochondria. Importantly activation of RhoA in cardiomyocytes confers protection against ischemia, and this cardioprotection is attenuated by PINK1 knockdown. In vivo LAD ligation elicits increases in mitochondrial PINK1, Parkin, and ubiquitinated mitochondrial proteins in the heart. AAV9-mediated RhoA expression potentiates these responses and a concurrent decrease in infarct size is observed. Remarkably, induction of mitochondrial PINK1 accumulation in response to RhoA signaling is neither mediated through its transcriptional upregulation nor dependent on mitochondrial membrane depolarization. Instead, RhoA signaling inhibits PINK1 cleavage, thereby stabilizing PINK1 protein at mitochondria. We further show that active RhoA interacts with PINK1 at mitochondria, and that this is regulated by RhoA downstream effector protein kinase D. These findings demonstrate for the first time that RhoA activation engages a unique mechanism to induce mitophagy and protect the heart against ischemic stress, and implicates regulation of RhoA signaling as a potential strategy to provide cardioprotection via regulation of mitophagy.