Mitochondria are dynamic organelles, undergoing structural and functional changes in response to cellular needs such as metabolic demands, calcium signaling, and apoptosis. Given its location at the interface of mitochondria and other cellular compartments, the mitochondrial outer membrane (MOM) voltage-dependent anion channel (VDAC) serves as a cellular hub for the dynamic regulation of mitochondrial function. In vitro, VDAC switches from anionic ATP-permeable state to cationic calcium-favorable state under applied voltage and through voltage-dependent complexation with α-synuclein and tubulin. However, the presence of potential across MOM in live cells remains controversial. Hexokinase (HK), the rate-limiting enzyme in glycolysis, has been suggested to complex with VDAC to generate MOM potential (ΔΨ MOM ). Our results demonstrate metabolism-dependent generation and regulation of ΔΨ MOM (∼24 mV) and, thereby, potential voltage regulation of VDAC in HeLa cells through HK-VDAC complexation. HEK293 cells lacked appreciable ΔΨ MOM . This may be due to overexpression of HK2 in cancer cells, which has been linked to increased glycolysis (Warburg effect). Using HeLa cells with CRISPR-Cas9 knockout (KO) of individual VDAC isoforms (VDAC1, 2, and 3), we demonstrate that VDAC1 specifically regulates glycolysis through its interaction with HK. Though there is no change in mitochondrial respiration, VDAC1KO decreases mitochondrial network complexity and cristae density, while VDAC2KO, which is lethal in mouse models, has no effect on mitochondrial respiration despite upregulation of electron transport chain complexes. There is an increased cristae density in VDAC2KO, linked to decreased mitochondrial calcium uptake from the endoplasmic reticulum. Surprisingly, VDAC3KO decreased respiration and increased glutamine metabolism in mitochondria, suggesting that VDAC3, the oldest and minor isoform, is vital for mitochondrial metabolism. Our results highlight the complex regulation of MOM permeability, mitochondrial function, and cellular metabolism through dynamic isoform-specific complexation of VDAC with cytosolic proteins.
Lafargue et al. (Sun,) studied this question.