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Protocol for fluorescence-activated mitochondria sorting in endoplasmic reticulum-mitochondria contact reporter cells
Drew C Stark1, Brandon Chen2, Kayla Hickman3
1Cancer Biology Program, University of Michigan Medical School, Ann Arbor, MI 48109, USA; Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor, MI 48109, USA.
Abstract:
Mitochondria exchange metabolites bidirectionally with the endoplasmic reticulum to maintain bioenergetic homeostasis and respond to cellular stress. Here, we describe a protocol for generating cells that contain a split-GFP contact-site reporter of ER-mitochondria interactions and the sorting of heterogeneous mitochondrial populations based on fluorescence intensity. This approach enables reproducible functional profiling of mitochondrial subpopulations, validated by proteomic analysis demonstrating the enrichment of endoplasmic reticulum-associated proteins in GFP-positive mitochondria. For complete details on the use and execution of this protocol, please refer to Chen et al.1.