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Yeast Mitochondrial Derived Vesicles (MDVs) have membrane potential and contain a functional ATP synthase-Part 2

ABSTRACT: Mitochondria are responsible for the production of energy and for essential metabolic pathways in the cell, as well as fundamental processes such as apoptosis and aging. Interestingly, while the import machinery of proteins into mitochondria is thoroughly understood, there is currently no known protein export mechanism from mitochondria. Vesicular transport is a means of inter-organellar communication, and in fact cells release vesicles in order to shuttle lipids, proteins, RNA and DNA between one another. Here we show for the first time that functional mitochondria, isolated from Saccharomyces cerevisiae can release vesicles, independent of the fission machinery. Mitochondrial secreted vesicles are relatively uniform in size, of about 100nm and carry a selection of cargo; only certain proteins are carried by these vesicles, of which some are matrix soluble and some are membrane proteins. We further demonstrate that yeast MDVs harbor a membrane potential, a functional ATP synthase complex, produce ATP, and fuse with naïve mitochondria. We suggest, that MDVs may have a function in organellar protein export, ATP production ability of damaged mitochondria and organelle-communication.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Tamar Ziv

LAB HEAD: Pines Ophry

PROVIDER: PXD039191 | Pride | 2023-01-29

REPOSITORIES: Pride

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Project description:Mitochondria share attributes of vesicular transport with their bacterial ancestors given their ability to form mitochondrial-derived vesicles (MDVs). MDV formation enhances with stress and has been explored for its potential role in mitochondrial-directed cellular and extracellular signalling. Although the fate of MDVs as a form of mitochondrial quality control and cellular communication has been suggested, the proteomic cargo has remained mostly undefined. In this study, we strategically used an in vitro MDV budding/reconstitution assay, following graded oxidative stress on cardiac mitochondria, to identify and characterize the MDV proteome. Our results confirmed previously identified cardiac MDV markers while also revealing a complete map of the MDV proteome, paving the way to understanding the likely roles for MDV release. MDV cargo was particularly enriched with mitochondrial matrix and inner membrane proteins. Oxidative stress vulnerability of proteins directed the cargo loading, which was enhanced by antimycin A (Ant-A). Among OXPHOS complexes, complexes III and V were found to be Ant-A-sensitive. Proteins from metabolic pathways such as TCA cycle, amino acid, fatty acid and aldehyde metabolism, Fe-S cluster, antioxidant response proteins, autophagy and vesicular processes were also found to be Ant-A sensitive. Intriguingly, proteins containing hyper-reactive cysteine residues (HRCR), which act as redox switches in regulating metabolic pathways, including professional redox enzymes and those that are involved in iron metabolism were found to be loaded in MDVs with Ant-A sensitivity. Lastly, we estimated a possible significant contribution of MDVs to the formation of extracellular vesicles, which can serve as an indicator of mitochondrial stress. In conclusion, using graded oxidative stress, our study gives a layout of MDV proteomics signature, delineates the cargo selectivity and hints at the potential of MDVs and the novel protein cargo identified to serve as vital biomarkers during mitochondrial stress and its related pathologies.

Dataset Information

Yeast Mitochondrial Derived Vesicles (MDVs) have membrane potential and contain a functional ATP synthase-Part 2

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