However, it fails to reverse CCCP-induced AMPK activation, suggesting that CCCP-induced autophagy is dependent on ISR and independent of AMPK activation. ISRIB, an inhibitor of the ISR pathway, mitigates CCCP-induced expression of ATF4 and CHOP, activation of AKT, and autophagy, similar to NAC. All these events are neutralized by NAC, an anti-ROS, suggesting that CCCP-induced mitochondrial dysfunction promotes oxidative stress. CCCP also downregulates translation and promotes autophagy, leading to noncaspase-mediated cell death in HepG2 cells. Also, CCCP activates AMPK, a cellular energy sensor, and AKT, a regulator implicated in cell survival, and suppresses phosphorylation of mTORC1 substrates eIF4E-BP1 and S6K. Our findings suggest that the CCCP-induced ISR pathway is mediated by activation of HRI kinase, but not by GCN2, PERK, or PKR.
All rights reserved.Mitochondrial dysfunction mediated by CCCP (carbonyl cyanide m-chlorophenyl hydrazone), an inhibitor of mitochondrial oxidative phosphorylation, evokes the integrated stress response (ISR), which is analyzed here by eIF2α phosphorylation and expression profiles of ATF4 and CHOP proteins. In particular, we analyze the cellular modifications produced by the activation of two major pathways involving the signaling of the nuclear factor erythroid 2-related factor 2 (Nrf2) and the transcription factor EB (TFEB), and discuss the contribution of these pathways to the integrated cellular stress response.ĬCCP CCCP, Carbonyl Cyanide m Chlorophenyl Hydrazone (PubChem CID: 2603) FCCP, Carbonyl Cyanide p Trifluoromethoxyphenylhydrazone (PubChem CID: 3330) Mitochondria Mitophagy N-Acetyl-l-cysteine (PubChem CID: 12035) NRF2 TFEB.Ĭopyright © 2018 Elsevier Inc.
We here review current mechanistic insights into the induction of mitophagy and autophagy by CCCP and FCCP. Sustained perturbation of ΔΨm, which is normally tightly controlled to ensure cell proliferation and survival, triggers various stress pathways as part of the cellular adaptive response, the main components of which are mitophagy and autophagy. CCCP and FCCP are now being increasingly used for investigating the mechanisms of autophagy by inducing mitochondrial degradation through the disruption of the mitochondrial membrane potential (ΔΨm). The ring-substituted derivatives of carbonyl cyanide phenylhydrazone, CCCP and FCCP, are routinely used for the analysis of the mitochondrial function in living cells, tissues, and isolated mitochondrial preparations.
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