Ultrastructural Characterization of Flashing Mitochondria

Area of Science:

• Cell Biology
• Mitochondrial Biology
• Biophysics

Background:

• Mitochondria exhibit spontaneous transient matrix pH elevations (mitopHlashes) linked to membrane potential drops.
• The precise mechanism underlying mitopHlashes remains unclear, despite their dependence on the respiratory chain and optic atrophy 1 protein.

Purpose of the Study:

• To elucidate the structural basis of mitopHlashes by examining the ultrastructure of flashing mitochondria.
• To investigate the relationship between mitochondrial morphology and the occurrence of mitopHlashes.

Main Methods:

• Correlative light and electron microscopy (CLEM) was employed on HeLa cells expressing a matrix pH probe (mitoSypHer).
• Serial block face scanning electron microscopy (SBF-SEM) with focused ion beam (FIB) milling was used for high-resolution 3D ultrastructural reconstruction.
• Live fluorescence imaging was correlated with fixed electron microscopy data to identify flashing and nonflashing mitochondria.

Main Results:

• Flashing mitochondria were significantly larger (10-fold) than nonflashing ones, yet maintained similar surface-to-volume and cristae volumes.
• Flashing mitochondria exhibited increased membrane contact sites and were interconnected by tubular structures.
• A constriction was observed at a junction between flashing and nonflashing mitochondria, suggesting fusion pore dynamics.

Conclusions:

• MitopHlashes are associated with distinct mitochondrial morphology, characterized by increased size and interconnections.
• The structural findings support the hypothesis that mitopHlashes may be initiated by the opening of fusion pores between adjacent mitochondria.
• Flashing mitochondria are structurally intact and bioenergetically functional, despite their dynamic pH changes.