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Updated: Sep 13, 2025
Visualization of ATP Synthase Dimers in Mitochondria by Electron Cryo-tomography
Published on: September 14, 2014
In situ cryo-ET visualization of mitochondrial depolarization and mitophagic engulfment
Kevin Rose1,2,3, Eric Herrmann1,2,3, Eve Kakudji1,2,3
1Aligning Science Across Parkinson's Collaborative Research Network, Chevy Chase, MD 20815.
Abstract:
Defective mitochondrial quality control in response to loss of mitochondrial membrane polarization is implicated in Parkinson's disease by mutations in PINK1 and PRKN. Parkin-expressing U2 osteosarcoma (U2OS) cells were treated with the depolarizing agents oligomycin and antimycin A (OA) and subjected to cryo-focused ion beam milling and in situ cryo-electron tomography. Mitochondria were fragmented and devoid of matrix calcium phosphate crystals. Phagophores were visualized, with bridge-like lipid transporter densities connected to mitophagic phagophores. A subpopulation of ATP synthases relocalized from cristae to the inner boundary membrane. The structure of the dome-shaped prohibitin complex, a dodecamer of PHB1-PHB2 dimers, was determined in situ by subtomogram averaging in untreated and treated cells and found to exist in open and closed conformations, with the closed conformation being enriched by OA treatment. These findings provide a set of native snapshots of the manifold nano-structural consequences of mitochondrial depolarization and provide a baseline for future in situ dissection of Parkin-dependent mitophagy.
Insights
Mitochondrial depolarization triggers fragmentation and structural changes, revealing insights into Parkinson's disease mechanisms. This study visualizes key events in mitophagy, offering a baseline for future research on Parkin-dependent processes.
Area of Science:
- Cell Biology
- Neuroscience
- Biochemistry
Background:
- Defective mitochondrial quality control is linked to Parkinson's disease, particularly mutations in PINK1 and Parkin (PRKN).
- Loss of mitochondrial membrane potential is a critical trigger for mitophagy, a cellular process for removing damaged mitochondria.
Purpose of the Study:
- To investigate the nano-structural consequences of mitochondrial depolarization in situ.
- To visualize the early stages of Parkin-dependent mitophagy.
Main Methods:
- Utilized cryo-focused ion beam milling and in situ cryo-electron tomography on Parkin-expressing U2 osteosarcoma cells treated with depolarizing agents.
- Performed subtomogram averaging to determine the structure of the prohibitin complex in situ.
Main Results:
- Mitochondria showed fragmentation and loss of calcium phosphate crystals upon depolarization.
- Observed phagophores connected to mitochondria, with evidence of lipid transporter involvement.
- ATP synthases relocated from cristae to the inner boundary membrane.
- The prohibitin complex adopted distinct open and closed conformations, with the closed form enriched after treatment.
Conclusions:
- Mitochondrial depolarization induces significant nano-structural alterations.
- Provides in situ structural data on mitophagy initiation, serving as a foundation for studying Parkin-dependent mitophagy.
