Mitochondrial ROS-dependent localization and trafficking of glycolytic enzymes to mitochondria

ABSTRACT: Mitochondrial reactive oxygen species (mtROS) regulate cellular signaling pathways, but also cause oxidative stress when de-regulated during aging and pathological conditions such as neurodegenerative diseases. The dynamic trafficking of proteins between cellular compartments is a common mechanism to control their stability and biological activities. By targeting the BirA* biotin ligase to the outer mitochondrial membrane in HEK293 cells, we identified proteins whose labelling increased or decreased in response to menadione consistent with a change in their trafficking to and from mitochondria in response to increased ROS production. These proteins represent potential candidates for future studies of mitochondrial stress signaling. A subset of glycolytic enzymes was found in this screen and confirmed, by mitochondrial fractionation and imaging, to increase localization to mitochondria in response to menadione, despite no change in their overall abundance. Submitochondrial fractionation studies are consistent with localization of a pool of these enzymes to the mitochondrial intermembrane space. Localization of glycolytic enzymes to mitochondria was also increased in cells grown under hypoxia or that express a mitochondria-targeted D-amino acid oxidase (conditions that induce increased mtROS production), and inhibited basally under normal growth conditions by the mitochondrial antioxidant MnTBAP. Finally, primary Alzheimer’s disease (AD) fibroblasts also had glycolytic enzymes associated with mitochondria that was reversed by antioxidants, consistent with increased mtROS in these cells altering their relative cellular distribution. We speculate that the increased mitochondrial localization of glycolytic enzymes is an adaptive response to mtROS that alters glucose flux toward the antioxidant pentose phosphate pathway, creates distinct regulatory pools of mitochondrial metabolites or new metabolic circuits, and/or provides cytoprotection or other adaptive responses via moonlighting functions unrelated to their enzymatic activity.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Christian Münch  

LAB HEAD: Christian Münch

PROVIDER: PXD066204 | Pride | 2025-08-12

REPOSITORIES: Pride