Cytosolic translational responses differ under conditions of severe short-term and long-term mitochondrial stress

Abstract

Previous studies demonstrated that cells inhibit protein synthesis as a compensatory mechanism for mitochondrial dysfunction. Protein synthesis can be attenuated by 1) the inhibition of mTOR kinase, which results in a decrease in the phosphorylation of S6K1 and 4E-BP1 proteins, and 2) an increase in the phosphorylation of eIF2α protein. The present study investigated both of these pathways under conditions of short-term acute and long-term mitochondrial stress. Short-term responses were triggered in mammalian cells by treatment with menadione, antimycin A, or CCCP. Long-term mitochondrial stress was induced by prolonged treatment with menadione or rotenone and expression of genetic alterations, such as knocking down the MIA40 oxidoreductase or knocking out NDUFA11 protein. Short-term menadione, antimycin A, or CCCP cell treatment led to the inhibition of protein synthesis, accompanied by a decrease in mTOR kinase activity, an increase in the phosphorylation of eIF2α (Ser51), and an increase in the level of ATF4 transcription factor. Conversely, long-term stress led to a decrease in eIF2α (Ser51) phosphorylation and ATF4 expression and to an increase in S6K1 (Thr389) phosphorylation. Thus, under long-term mitochondrial stress, cells trigger long-lasting adaptive responses for protection against excessive inhibition of protein synthesis.

CCCP‐induced mitochondrial dysfunction – characterization and analysis of integrated stress response to cellular signaling and homeostasis

“…Our findings are consistent with some of the very recent observations where mitochondrial dysfunction caused by CCCP or oligomycin treatments are shown to promote the ISR pathway through the activation of HRI kinase [41,42]. Other studies have shown that mitochondrial dysfunction can evoke the ISR pathway but have not characterized the eIF2a kinase [54] or observed that it is not one of the four well-characterized kinases [59], and others have implicated GCN2 [60] or PERK [61]. HRI is an important hemoprotein in erythroid precursors that senses heme levels and is activated in the absence of heme, denatured proteins, oxidative stress, and regulated by molecular chaperones Hsp90 and Hsc70 [38,48].…”

“…We used a typical antioxidant, NAC, and recently discovered ISRIB, an ISR inhibitor to analyze the importance of CCCP-induced mitochondrial ROS as reported by others [53,54] in regulating the cellular signaling pathways as studied above and the cross talk between ISR pathway to cellular signaling and autophagy. HepG2 cells were pretreated with 5 mM NAC or with 250 nM ISRIB for 1 h before the addition of CCCP and incubated for different time points as mentioned in the figures.…”

4E-BP1 Protects Neurons from Misfolded Protein Stress and Parkinson's Disease Toxicity by Inducing the Mitochondrial Unfolded Protein Response

“…However, beyond this classical model of UPR mt activation, further research has demonstrated that the UPR mt is activated in response to a diverse array of both mitochondrial and cellular stresses. Indeed, recent studies have shown that activation of the integrated stress response, which turns off global protein translation on regulatory phosphorylation of eIF2a, culminates with activation of the UPR mt (Fiorese et al, 2016;Munch and Harper, 2016;Quiros et al, 2017;Samluk et al, 2019). To test whether 4E-BP1 overexpression, which also reduces global protein synthesis, could be activating the UPR mt , we measured the transcription levels of two master UPR mt regulators, CHOP and ATF4, whose transcriptional upregulation has been shown to yield UPR mt activation (Zhao et al, 2002;Wang et al, 2018).…”

Mitochondria–Lysosome Crosstalk: From Physiology to Neurodegeneration

“…The behavior of AMPK and mTORC1 in response to mitochondrial and lysosomal defects has predictable downstream implications given the importance of these regulatory nodes in cell functions. For example, protein synthesis is under tight regulation by mTORC1, and is strongly inhibited in acute mitochondrial stress, but is activated in chronic mitochondrial stress [85]. It remains unclear whether these effects are related to changes in AMPK; however, the fact that mTORC1 and translation are inhibited when AMPK is hyperactive (acute mitochondrial stress), and active when AMPK is repressed [14,17], suggests that AMPK might be involved.…”