Fasting Depletes a Key Metabolite That Triggers Mitochondrial Cleanup

Mitophagy — the selective autophagic clearance of dysfunctional mitochondria — is fundamental to cellular quality control and longevity. While the PINK1–Parkin pathway is well characterized, less is known about how nutritional status, particularly fasting, activates mitophagy through receptor-mediated pathways. This Nature paper by Zhang, Shen, Shen, Wang et al. identifies cytosolic acetyl-coenzyme A (AcCoA) as a direct metabolic signal linking nutrient availability to mitophagy, mediated by the mitochondrial outer membrane protein NLRX1.

The researchers began by modeling fasting conditions in vitro using a mild starvation medium (SM: 5 mM glucose, 2 mM glutamine), mirroring serum metabolite changes observed in overnight-fasted mice. SM induced measurable mitophagy — demonstrated by elevated acidic mt-Keima reporter signals, decreased mtDNA/nDNA ratios, and reduced mitochondrial proteins TIM23, MT-CO2, and HSP60 — in HeLa, A549, and MCF7 cells, but not in U-2 OS cells. This mitophagy was blocked by bafilomycin A1, confirming autophagic flux. Crucially, SM did not activate AMPK (no increase in p-AMPK or p-ULK1-S555) and did not suppress mTORC1, distinguishing this pathway from canonical starvation-induced autophagy.

Mass spectrometry revealed that cytosolic — but not mitochondrial — AcCoA levels specifically decreased in SM-responsive cells (HeLa, A549, MCF7), not in the non-responsive U-2 OS cells. These responsive cells also had significantly higher baseline expression of ACLY, FASN, IDH1, SLC25A1, and ACC1 — all enzymes involved in cytosolic AcCoA metabolism. Pharmacological inhibition of ACLY (using HC or SB204990) or SLC25A1 (using BTC) reduced cytosolic AcCoA and enhanced mitophagy. Genetic knockdown of ACLY, SLC25A1, or ACSS2 replicated these effects in normal medium. Acetate supplementation — which replenishes cytosolic AcCoA via ACSS2 — abolished SM-induced and ACLY-knockdown-induced mitophagy, confirming the causal role of cytosolic AcCoA.

To identify the molecular mediator, the team performed a genome-wide CRISPR screen for genes required for HC-induced mitophagy. NLRX1 emerged as the top-ranked mitophagy receptor gene. Knockout of NLRX1 abolished mitophagy induced by SM, ACLY inhibition, or SLC25A1 inhibition both in cell culture and in vivo. In Nlrx1−/− mice, intraperitoneal HC injection failed to reduce mitochondrial mass in liver tissue, and mt-Keima signals confirmed absent mitophagy response compared with wild-type controls.

Mechanistically, structural and biochemical analyses showed that cytosolic AcCoA binds directly to a conserved pocket within NLRX1's leucine-rich repeat (LRR) domain. This binding stabilizes an intramolecular interaction between the LRR and NACHT domains, maintaining NLRX1 in an autoinhibited conformation that prevents its association with the autophagy adaptor LC3. When AcCoA levels fall, this autoinhibition is relieved, NLRX1 binds LC3, and mitophagy proceeds. Finally, the study found that KRAS inhibitor treatment (which suppresses glycolysis and reduces cytosolic AcCoA) activates NLRX1-dependent mitophagy, and that this response promotes cancer cell survival and drug resistance — nominating the AcCoA–NLRX1 axis as a therapeutic target to overcome KRAS inhibitor resistance in cancer.