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Cell-Free Analysis of Mitochondrial Fusion by Fluorescence Microscopy

  • Nyssa Becker Samanas
  • Suzanne HoppinsEmail author
Protocol
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Part of the Methods in Molecular Biology book series (MIMB, volume 2159)

Abstract

Dynamin-related proteins on both the mitochondrial outer and inner membranes mediate membrane fusion. Mitochondrial fusion is regulated in many different physiological contexts including cell cycle progression, differentiation pathways, stress responses, and cell death. Mitochondrial fusion is opposed by mitochondrial division and requires movement of mitochondria on microtubules. We developed a cell-free reconstituted mitochondrial fusion assay to circumvent the complexity of the pathways impinging on the activity of the mitochondrial fusion machinery in vivo. This allows for quantification of mitochondrial fusion in defined conditions and in the absence of other processes such as mitochondrial division or transport. The impact of proteins or small molecules on mitochondria fusion can also be assessed. Here we describe the cell-free mitochondrial fusion assay using mitochondria isolated from mouse embryonic fibroblasts.

Key words

Mitochondrial dynamics Fusion Cell-free Isolated mitochondria Mitofusin Opa1 Microscopy 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  1. 1.Department of BiochemistryUniversity of WashingtonSeattleUSA

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