The Role of GRAF RHOA-GAPs in Maintaining Highly Energetic Tissues via Regulation of Mitochondrial Quality Control

Mitochondrial quality control is a biological process of critical importance for the maintenance of cellular function and viability, especially in highly energetic, terminally differentiated tissues such as skeletal muscles and the brain. A key aspect of mitochondrial quality control is the degradation of damaged mitochondria via the autophagic process of mitophagy. In this dissertation, we explore novel roles for two related RHOA GTPase activating proteins, GRAF2 and GRAF1, in maintaining the health of skeletal muscle and brain, respectively, via regulation of PARKIN-dependent mitophagy. We further detail the development of an automated image analysis pipeline for the study of striated muscle biology which was of great utility in these and other studies which we have performed.In skeletal muscle, mitophagy is necessary for baseline maintenance of the mitochondrial pool as well as for adaptive responses to exercise. Herein, we demonstrate that GRAF2 is highly expressed throughout adulthood in mouse skeletal muscles and characterize a novel skeletal muscle-restricted GRAF2 knockout mouse model. Functionally, we find that loss of GRAF2 results in a dramatic reduction in endurance exercise capacity. This dysfunction is shown to be underpinned by defects in the regulation of oxidative metabolism as assessed by transcriptomic, metabolomic, and muscle fiber type composition analyses. In investigating the molecular mechanism of these effects, we uncover a novel role for GRAF2 in the regulation of mitophagy as a facilitator of PARKIN/LC3 complex formation, actin remodeling, and retrograde mitochondrial trafficking. Mitophagy is also crucial for neuronal health, and defects in the brain are known to play a pathogenic role in Parkinson’s disease. Through the preliminary studies detailed in this dissertation, we show that there is a genetic interaction between GRAF1 and PARKIN in the brains of a preclinical mouse model of Parkinson’s disease. Specifically, we demonstrate a loss of dopaminergic neurons in the substantia nigra accompanied by behavioral changes consistent with the early onset of Parkinson’s disease. Together, these studies establish an exciting new role for GRAFs in skeletal muscle and brain health via mitophagy regulation.