Mitochondrial Fusion In Vitro

Abstract

The field of mitochondrial dynamics has received a great deal of attention as a result of a number of studies linking mitochondrial fission and fusion machinery to apoptosis. Specifically, elevated levels of mitochondrial fission or compromised mitochondrial fusion can sensitize cells to apoptotic stimuli. Conversely, stimulation of mitochondrial fusion can render cells resistant to apoptotic stimuli. In addition, the machinery involved in fission and fusion has been spatially linked to Bax, a pro-apoptotic protein. However, the mechanistic implications of interactions between the machinery of mitochondrial fission and fusion and apoptotic effectors are largely unknown. Our understanding of the pathways of mitochondrial fission and fusion have come from genetic studies coupled with direct observation of both fission and fusion components and mitochondrial organelle morphology and behavior in vivo in Saccharomyces cerevisiae. These approaches have identified the key players in both mitochondrial fission and fusion and have generated good models for their roles in mitochondrial dynamics. However, the lack of in vitro systems for studying these processes has impeded a deeper investigation of the mechanism. We have recapitulated the process of mitochondrial fusion in vitro (5). Using this in vitro fusion assay, we have separated outer mitochondrial membrane fusion from inner and identified the mechanistic requirements for each step.