Characterizing the molecular mechanisms of mitochondrial fusion and division in healthy and diseased cells

Abstract

Balanced mitochondrial fusion and division activity in healthy cells typically establishes a reticular mitochondrial network that is able to support normal cellular activities. Disruptions in balanced mitochondrial dynamics can occur through regulated changes in fusion and division to mediate physiological processes such as cell division and apoptosis. Additionally, imbalanced mitochondrial dynamics have been reported as both a cause and consequence of several neurodegenerative diseases. I begin with a discussion of imbalanced mitochondrial dynamics in disease and the therapeutic potential of drugs that target the mitochondrial fusion and division machines. While the causative role of mitochondrial dynamics has only been established in a subset of diseases, I discuss our current understanding of how aberrant mitochondrial dynamics influences neurodegenerative, cardiac and metabolic diseases. I next discuss published work in which I characterized four mutations in the gene DNM1L, which encodes the mitochondrial division protein Drp1. All mutations were identified by whole exome sequencing in patients with neurological defects and mitochondrial dysfunction. To determine if the mutations were sufficient to cause mitochondrial hyperfusion, I expressed each of the four mutations in human and yeast cells lacking Drp1. I also evaluated mitochondrial morphology in wild type human and yeast cells expressing each mutant to determine if the division defects were dominant as is observed in human disease. A novel mutation in the GTPase domain of Drp1, G32A, was of particular interest as a compelling causal mutation that inhibits Drp1 recruitment to mitochondria. Finally, I investigated the mechanism of mitochondrial fusion using phosphorylation mimicking and blocking mutations at a novel Mfn1 phosphorylation site, S228. My work suggests that phosphorylation of Mfn1 S228 inhibits mitochondrial fusion by inhibiting trans Mfn1 interactions and nucleotide-dependent Mfn1 assembly. I also describe in vivo mitochondrial clustering and hyperfusion in cells expressing Mfn1 S228E and discuss a model to reconcile differences between cellular phenotypes and biochemical fusion defects. This work will also be foundational to understand how one or more signaling pathways can influence Mfn1 S228 phosphorylation to coordinate mitochondrial morphology with cellular conditions. Together, my work has provided important insights into the relationship between mitochondrial dynamics and disease, specifically DNM1L-related neurodegeneration, as well as uncovered a possible regulatory mechanism to alter mitofusin activity based on cellular conditions.