FIG. 8.

Inhibition of the malate/aspartate NADH shuttle induces reductive stress and metabolic reprogramming. (A) The malate/aspartate shuttle exchanges cytosolic NAD(H) with mitochondrial NAD(H) to maintain high cytosolic NAD⁺ levels, which are required for glycolysis, and high mitochondrial NADH levels, which provide electrons for mitochondrial OXPHOS. (B) Inhibition of the malate/aspartate shuttle by silencing GOT2, suppressing SIRT3 activity, or using the chemical inhibitor AOA leads to accumulation of cytosolic NADH and increases in cytosolic NADH/NAD⁺ and reductive ROS generation, indicative of reductive stress. NADH-induced reductive stress shifts cell metabolism from mitochondrial respiration to glycolysis. Enhancement of this shuttle activity by overexpression of AGC1 or recovery of GOT2 activity significantly increases mitochondrial NADH levels and enhances mitochondrial respiration. AGC1, aspartate-glutamate carrier 1; AOA, aminooxyacetic acid; GOT2, glutamate-oxaloacetate transaminase; Mal-Asp shuttle, malate/aspartate shuttle; OXPHOS, oxidative phosphorylation; SIRT3, sirtuin deacetylase family member 3.