Abstract
Mitochondrial flashes mediated by optic atrophy 1 (OPA1) fusion protein are bioenergetic responses to stochastic drops in mitochondrial membrane potential (Δψm) whose origin is unclear. Using structurally distinct genetically encoded pH‐sensitive probes, we confirm that flashes are matrix alkalinization transients, thereby establishing the pH nature of these events, which we renamed “mitopHlashes”. Probes located in cristae or intermembrane space as verified by electron microscopy do not report pH changes during Δψm drops or respiratory chain inhibition. Opa1 ablation does not alter Δψm fluctuations but drastically decreases the efficiency of mitopHlash/Δψm coupling, which is restored by re‐expressing fusion‐deficient OPA1K301A and preserved in cells lacking the outer‐membrane fusion proteins MFN1/2 or the OPA1 proteases OMA1 and YME1L, indicating that mitochondrial membrane fusion and OPA1 proteolytic processing are dispensable. pH/Δψm uncoupling occurs early during staurosporine‐induced apoptosis and is mitigated by OPA1 overexpression, suggesting that OPA1 maintains mitopHlash competence during stress conditions. We propose that OPA1 stabilizes respiratory chain supercomplexes in a conformation that enables respiring mitochondria to compensate a drop in Δψm by an explosive matrix pH flash.
Highlights
Proton transfer across the inner membrane of mitochondria (IMM) couples the oxidation of carbohydrates and fat to the synthesis of adenosine triphosphate (ATP) that cells use as an energy source
optic atrophy 1 (OPA1) cleavage leads to the production of short, soluble forms of OPA1 (S-OPA1) that interact with uncleaved forms of OPA1 (L-OPA1) at cristae junctions and with subunits of the mitochondrial contact site (MICOS) complex involved in the maintenance of cristae structure [24,25,26]
We further show that OPA1 is not required for stochastic Dwm fluctuations but allows efficient generation of mitopHlashes, that L-OPA1 isoforms can restore flashing independently of mitochondrial fusion, and that loss of OPA1-mediated mitopHlash/Dwm coupling is an early marker of apoptosis
Summary
Proton transfer across the inner membrane of mitochondria (IMM) couples the oxidation of carbohydrates and fat to the synthesis of adenosine triphosphate (ATP) that cells use as an energy source. The OPA1 gene is alternatively spliced at exons 4, 4a, and 5b leading to the expression of eight variants that differ in their ability to trigger membrane fusion and in their resistance to apoptosis [18,19,20]. These variants can be constitutively cleaved by the IMM peptidase OMA1 and the i-AAA protease YME1L at sites located in exon 5 and 5b, respectively [21,22,23]. Recent findings suggest that S-OPA1 is involved in IMM fission [27]
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