Oxidation of Alpha-Ketoglutarate Is Required for Reductive Carboxylation in Cancer Cells with Mitochondrial Defects

Abstract

Summary Mammalian cells generate citrate by decarboxylating pyruvate in the mitochondria to supply the tricarboxylic acid (TCA) cycle. In contrast, hypoxia and other impairments of mitochondrial function induce an alternative pathway that produces citrate by reductively carboxylating α-ketoglutarate (AKG) via NADPH-dependent isocitrate dehydrogenase (IDH). It is unknown how cells generate reducing equivalents necessary to supply reductive carboxylation in the setting of mitochondrial impairment. Here we identified shared metabolic features in cells using reductive carboxylation. Paradoxically, reductive carboxylation was accompanied by concomitant AKG oxidation in the TCA cycle. Inhibiting AKG oxidation decreased reducing equivalent availability and suppressed reductive carboxylation. Interrupting transfer of reducing equivalents from NADH to NADPH by nicotinamide nucleotide transhydrogenase increased NADH abundance and decreased NADPH abundance while suppressing reductive carboxylation. The data demonstrate that reductive carboxylation requires bidirectional AKG metabolism along oxidative and reductive pathways, with the oxidative pathway producing reducing equivalents used to operate IDH in reverse.

L-2-Hydroxyglutarate production arises from noncanonical enzyme function at acidic pH

“…Acidic pH appears to enhance LDHA-mediated reduction of αKG by driving equilibrium toward the protonated state of the carboxylate tail, which permits docking of αKG via interaction with the glutamine 100 in LDHA. Intracellular acidification might also explain, at least in part, the increased L-2HG observed in cells with electron transport chain dysfunction 46 .…”

Mechanisms of redox metabolism and cancer cell survival during extracellular matrix detachment

“…For example, in order for cancer cells to survive glucose deprivation, glutamate dehydrogenase activity increases to support the synthesis of TCA cycle intermediates that can be utilized for macromolecular and nucleotide synthesis (36,37). More salient for the topic at hand, glutamine metabolism is critically important for the production of glutathione and NADPH which both function to detoxify ROS in a fashion that is required for cancer cell survival (19,(37)(38)(39)(40)(41)(42)(43)(44). Moreover, certain types of cancers appear to have a specific mechanism to utilize glutamine to effectuate their progression.…”

Compartmentation of Metabolites in Regulating Epigenomes of Cancer

“…Recent reports found that the level of FAD has an impact on the activity of LSD1. For instance, riboflavin deficiency in Jurkat increased α-KG was observed, which leads to a pan-cancer genome reduction of DNA methylation, especially at the highly repetitive sequences and lowdensity CpG regions (44,54,62,63,64). But a recent study actually suggested that hypoxia or lactate acidosis, both conditions commonly associated with almost all types of solid tumor growth, decrease α-KG, and at least one gene, ETV4, was identified as regulated by α-KG epigenetically (65).…”

“…Intracellular α-KG is derived mainly from the glycolysis pathway via decarboxylation of pyruvate and the TCA anaplerosis pathway via glutaminolysis (53). Simultaneously, α-KG is also converted to citrate, catalyzed by iso-citrate dehydrogenases (IDHs), and to succinate, catalyzed by α-KG dehydrogenase (53,54). Thus, intracellular enrichment of α-KG is very dynamic and subject to the cellular fluctuation of metabolic activities.…”