Dataset Information

Raptor couples mTORC1 and ERK1/2 inhibition by cardamonin with oxidative stress induction in ovarian cancer cells.

ABSTRACT:

Background

A balance on nutrient supply and redox homeostasis is required for cell survival, and increased antioxidant capacity of cancer cells may lead to chemotherapy failure.

Objective

To investigate the mechanism of anti-proliferation of cardamonin by inducing oxidative stress in ovarian cancer cells.

Methods

After 24 h of drug treatment, CCK8 kit and wound healing test were used to detect cell viability and migration ability, respectively, and the ROS levels were detected by flow cytometry. The differential protein expression after cardamonin administration was analyzed by proteomics, and the protein level was detected by Western blotting.

Results

Cardamonin inhibited the cell growth, which was related to ROS accumulation. Proteomic analysis suggested that MAPK pathway might be involved in cardamonin-induced oxidative stress. Western blotting showed that cardamonin decreased Raptor expression and the activity of mTORC1 and ERK1/2. Same results were observed in Raptor KO cells. Notably, in Raptor KO cells, the effect of cardamonin was weakened.

Conclusion

Raptor mediated the function of cardamonin on cellular redox homeostasis and cell proliferation through mTORC1 and ERK1/2 pathways.

SUBMITTER: Zhu Y

PROVIDER: S-EPMC10257395 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

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Publications

Raptor couples mTORC1 and ERK1/2 inhibition by cardamonin with oxidative stress induction in ovarian cancer cells.

Zhu Yanting Y Wang Shifeng S Niu Peiguang P Chen Huajiao H Zhou Jintuo J Jiang Li L Li Danyun D Shi Daohua D

PeerJ 20230607

<h4>Background</h4>A balance on nutrient supply and redox homeostasis is required for cell survival, and increased antioxidant capacity of cancer cells may lead to chemotherapy failure.<h4>Objective</h4>To investigate the mechanism of anti-proliferation of cardamonin by inducing oxidative stress in ovarian cancer cells.<h4>Methods</h4>After 24 h of drug treatment, CCK8 kit and wound healing test were used to detect cell viability and migration ability, respectively, and the ROS levels were detec ...[more]

PMID: 37304865

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Project description:BackgroundmTORC1 (mechanistic target of rapamycin complex 1) is associated with lymphoma progression. Oncogenic RRAGC (Rag guanosine triphosphatase C) mutations identified in patients with follicular lymphoma facilitate the interaction between Raptor (regulatory protein associated with mTOR) and Rag GTPase. It promotes the activation of mTORC1 and accelerates lymphomagenesis. Cardamonin inhibits mTORC1 by decreasing the protein level of Raptor. In the present study, we investigated the inhibitory effect and possible mechanism of action of cardamonin in RRAGC-mutant lymphoma. This could provide a precise targeted therapy for lymphoma with RRAGC mutations.MethodsCell viability was measured using a cell counting kit-8 (CCK-8) assay. Protein expression and phosphorylation levels were determined using western blotting. The interactions of mTOR and Raptor with RagC were determined by co-immunoprecipitation. Cells overexpressing RagC wild-type (RagCWT) and RagC Thr90Asn (RagCT90N) were generated by lentiviral infection. Raptor knockdown was performed by lentivirus-mediated shRNA transduction. The in vivo anti-tumour effect of cardamonin was assessed in a xenograft model.ResultsCardamonin disrupted mTOR complex interactions by decreasing Raptor protein levels. RagCT90N overexpression via lentiviral infection increased cell proliferation and mTORC1 activation. The viability and tumour growth rate of RagCT90N-mutant cells were more sensitive to cardamonin treatment than those of normal and RagCWT cells. Cardamonin also exhibited a stronger inhibitory effect on the phosphorylation of mTOR and p70 S6 kinase 1 in RagCT90N-mutant cells. Raptor knockdown abolishes the inhibitory effects of cardamonin on mTOR. An in vivo xenograft model demonstrated that the RagCT90N-mutant showed significantly higher sensitivity to cardamonin treatment.ConclusionsCardamonin exerts selective therapeutic effects on RagCT90N-mutant cells. Cardamonin can serve as a drug for individualised therapy for follicular lymphoma with RRAGC mutations.

Project description:BackgroundThe integrated stress response (ISR) is an evolutionarily conserved process to cope with intracellular and extracellular disturbances. Myocardial infarction is a leading cause of death worldwide. Coronary artery reperfusion, the most effective means to mitigate cardiac damage of myocardial infarction, causes additional reperfusion injury. This study aimed to investigate the role of the ISR in myocardial ischemia/reperfusion (I/R).MethodsCardiac-specific gain- and loss-of-function approaches for the ISR were used in vivo. Myocardial I/R was achieved by ligation of the cardiac left anterior descending artery for 45 minutes followed by reperfusion for different times. Cardiac function was assessed by echocardiography. Cultured H9c2 cells, primary rat cardiomyocytes, and mouse embryonic fibroblasts were used to dissect underlying molecular mechanisms. Tandem mass tag labeling and mass spectrometry was conducted to identify protein targets of the ISR. Pharmacologic means were tested to manipulate the ISR for therapeutic exploration.ResultsWe show that the PERK (PKR-like endoplasmic reticulum resident kinase)/eIF2α (α subunit of eukaryotic initiation factor 2) axis of the ISR is strongly induced by I/R in cardiomyocytes in vitro and in vivo. We further reveal a physiologic role of PERK/eIF2α signaling by showing that acute activation of PERK in the heart confers robust cardioprotection against reperfusion injury. In contrast, cardiac-specific deletion of PERK aggravates cardiac responses to reperfusion. Mechanistically, the ISR directly targets mitochondrial complexes through translational suppression. We identify NDUFAF2 (NADH:ubiquinone oxidoreductase complex assembly factor 2), an assembly factor of mitochondrial complex I, as a selective target of PERK. Overexpression of PERK suppresses the protein expression of NDUFAF2 and PERK inhibition causes an increase of NDUFAF2. Silencing of NDUFAF2 significantly rescues cardiac cell survival from PERK knockdown under I/R. We show that activation of PERK/eIF2α signaling reduces mitochondrial complex-derived reactive oxygen species and improves cardiac cell survival in response to I/R. Moreover, pharmacologic stimulation of the ISR protects the heart against reperfusion damage, even after the restoration of occluded coronary artery, highlighting clinical relevance for myocardial infarction treatment.ConclusionsThese results suggest that the ISR improves cell survival and mitigates reperfusion damage by selectively suppressing mitochondrial protein synthesis and reducing oxidative stress in the heart.

Dataset Information

Raptor couples mTORC1 and ERK1/2 inhibition by cardamonin with oxidative stress induction in ovarian cancer cells.

Background

Objective

Methods

Results

Conclusion

Publications

Raptor couples mTORC1 and ERK1/2 inhibition by cardamonin with oxidative stress induction in ovarian cancer cells.

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