Mitophagy, a mitochondria-specific form of autophagy, removes dysfunctional mitochondria and is hence an essential process contributing to mitochondrial quality control. PTEN-induced
kinase 1 (PINK1) and the
E3 ubiquitin ligase Parkin are critical molecules involved in stress-induced mitophagy, but the intracellular signaling mechanisms by which this pathway is regulated are unclear. We tested the hypothesis that signaling through RhoA, a
small GTPase, induces mitophagy via modulation of the PINK1/Parkin pathway as a protective mechanism against ischemic stress. We demonstrate that expression of constitutively active RhoA as well as
sphingosine-1-phosphate induced activation of endogenous RhoA in cardiomyocytes result in an accumulation of PINK1 at mitochondria. This is accompanied by translocation of Parkin to mitochondria and ubiquitination of
mitochondrial proteins leading to recognition of mitochondria by autophagosomes and their lysosomal degradation. Expression of RhoA in cardiomyocytes confers protection against
ischemia, and this cardioprotection is attenuated by
siRNA-mediated PINK1 knockdown. In vivo
myocardial infarction elicits increases in mitochondrial PINK1, Parkin, and ubiquitinated
mitochondrial proteins. AAV9-mediated RhoA expression potentiates these responses and a concurrent decrease in
infarct size is observed. Interestingly, induction of mitochondrial PINK1 accumulation in response to RhoA signaling is neither mediated through its transcriptional upregulation nor dependent on depolarization of the mitochondrial membrane, the canonical mechanism for PINK1 accumulation. Instead, our results reveal that RhoA signaling inhibits PINK1 cleavage, thereby stabilizing PINK1
protein at mitochondria. We further show that active RhoA localizes at mitochondria and interacts with PINK1, and that the mitochondrial localization of RhoA is regulated by its downstream effector
protein kinase D. These findings demonstrate that RhoA activation engages a unique mechanism to regulate PINK1 accumulation, induce mitophagy and protect against ischemic stress, and implicates regulation of RhoA signaling as a potential strategy to enhance mitophagy and confer protection under stress conditions.