ArticleMitochondrial communications2025
A chemical-genetic approach to target voltage-sensitive fluorophores to mitochondria.
Article in Mitochondrial communications, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Authors and funding
5 authors.
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Abstract
Mitochondria play central roles in the physiology of eukaryotic cells. Mitochondrial membrane potential, in turn, is a key driver of mitochondrial physiology. We previously developed a system to localize voltage-sensitive fluorophores to mitochondria based on the hydrolysis of labile acetoxymethyl (AM) esters. One potential problem with this system is the premature hydrolysis of the labile AM ester prior to accumulation in the mitochondria. A possible solution is to replace the AM ester with a bulky cyclopropylmethylacetoxy (CPM) ester, which resists uncatalyzed hydrolysis but can be removed by certain esterases. When paired with exogenous expression of mitochondrially-targeted esterases like porcine liver esterase (PLE), this chemical-genetic hybrid approach can improve localization to mitochondria. In this manuscript, we use superresolution microscopy to show that a variety of proteins, including esterases from pig and bacteria can be effectively localized to mitochondria. Further, we establish that a CPM-modified rhodamine voltage reporter (RhoVR-CPM) shows improved localization to mitochondria in cells expressing mitochondrially-targeted esterases. Finally, RhoVR-CPM can be paired with fluorescence lifetime imaging microscopy (FLIM) to map changes in mitochondrial membrane potential.
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