BRET-Based Mitochondrial Subcompartment Localization Biosensors
Abdulrahman, A.; Lescuyer, M.; El Zein, R.; Cecon, E.; Dam, J.; Masri, B.; Jockers, R.
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The last decade has witnessed a marked increase in interest in mitochondria, whose dysfunction leads to the development of multiple diseases. Mitochondria are unique as they are highly compartmentalized organelles that are composed of two closely apposed membranes and whose biological function relies on the precise localization of nuclear-encoded proteins in distinct mitochondrial subcompartments. Here we developed a series of bioluminescence resonance energy transfer (BRET)-based localization biosensors to monitor the precise localization of proteins in different mitochondrial subcompartments (outer and inner membrane, intermembrane space at the inner boundary membrane, crista lumen and matrix) with a high spatial resolution (1-10 nm). These biosensors detected the correct localization and orientation of TOM20, TOM22, VDAC1, MICU1, ATP5F1C, OTC, and SIRT2/3 proteins in their respective subcompartments, as well as the translocation of BAX and Drp1 from the cytosol to mitochondria in intact cells. The localization sensors provide non-invasive tools to monitor protein localization and translocation to mitochondria in real-time with nanometer resolution in intact cells submitted to various stressors.
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