The effects of mitochondrial damaging reagents on membrane potential and mitophagy induction, with implications in neurodegeneration

Abstract

Mitochondria are double membrane-bound organelles with established roles in metabolism, biosynthesis, and energy production. Various mitochondrial quality control mechanisms have evolved to maintain the mitochondrial network. One such mechanism is mitophagy, where damaged mitochondria are removed from the cell via autophagosome engulfment and lysosome degradation. Mutations and deficiencies in mitophagy components are linked to Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis (ALS), suggesting mitochondrial regulation is vital for cellular homeostasis. As a result, research efforts have focused on understanding mitophagy by inducing the pathway using various mitochondrial damaging reagents. However, these studies have resulted in discrepancies in the literature, highlighting our gap in knowledge of the type and severity of mitochondrial damage that induces mitophagy and drives neurodegeneration. Here, I used quantitative live-cell imaging of HeLa cells to determine whether four mitochondrial damaging reagents are comparable in depolarizing the mitochondrial membrane potential and inducing mitophagy. These reagents include carbonyl cyanide m-chlorophenyl hydrazone (CCCP), valinomycin, rotenone, and deferiprone (DFP). I found that valinomycin is the most potent at uncoupling the membrane potential, and that only reagents that depolarize the membrane induce mitophagy. My results shed light on comparable and effective in vitro mitophagy-inducing damaging paradigms that could be used to further our understanding of the etiology of neurodegenerative diseases and potential therapies.