scholarly journals Salubrinal Regulates the Apoptosis of Adrenocortical Carcinoma Cells via the PERK/eIF2α/ATF4 Signaling Pathway

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Lili Wu ◽  
Chunfeng Liang ◽  
Xuemei Huang ◽  
Xiujun Deng ◽  
Jiming Jiang ◽  
...  
Keyword(s):  
Cell Viability ◽  
Signaling Pathway ◽  
Adrenocortical Carcinoma ◽  
Cell Apoptosis ◽  
Translation Initiation Factor ◽  
Ion Concentration ◽  
Initiation Factor ◽  
Confocal Laser ◽  
Related Proteins

The protein-kinase-R- (PKR-) like endoplasmic reticulum kinase (PERK) signaling pathway is a well-known promoter of cell apoptosis. In this study, we aimed to determine whether salubrinal (Sal), a selective activator of eukaryotic translation initiation factor 2 (eIF2α), can induce apoptosis of human adrenocortical carcinoma (ACC) cell via activating the PERK/eIF2α/ATF4 signaling pathway, and the potential mechanisms of this action were explored. The ACC cell lines, including SW-13 and NCI–H295 R, were used. 3-(4,5)-Dimethylthiazol(-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, cell scratch experiments, flow cytometry, and JC-1 staining assays were performed to detect the cell viability, cell migration, and cell apoptosis. The expression of PERK/eIF2α/ATF4 signaling-pathway-related proteins and apoptosis-related proteins was detected by western blot (WB). Intracellular Ca2+ ion concentration was determined by a confocal laser scanning microscope. The results showed that Sal inhibited the migration and proliferation of ACC cells. Sal remarkably increased the influx of Ca2+ ion and the apoptosis rate of ACC cells in vitro. Furthermore, the expression levels of PERK/eIF2α/ATF4 signaling-related proteins and apoptosis-related proteins were upregulated in the treatment of Sal. The research demonstrated that Sal reduces the cell viability, increases the intracellular calcium concentration, and promotes the apoptosis of ACC cells in vitro through increasing the phosphorylation level of eIF2α and activating the PERK/eIF2α/ATF4 signaling. PERK/eIF2α/ATF4 is expected to act as a potential therapeutic target for the treatment of adrenocortical carcinoma.

scholarly journals Cannabinoid receptor 2 deletion deteriorates myocardial infarction through the down-regulation of AMPK-mTOR-p70S6K signaling-mediated autophagy

2019 ◽  
Vol 39 (4) ◽  
Author(s):  
Yao Hu ◽  
Yu Tao ◽  
Jing Hu
Keyword(s):  
Myocardial Infarction ◽  
Cell Viability ◽  
Signaling Pathway ◽  
Cannabinoid Receptor ◽  
Cell Counting ◽  
Protective Effects ◽  
Cannabinoid Receptor 2 ◽  
Related Proteins

AbstractCannabinoid receptor 2 (CB2R) has been reported to play an important role in the regulation of pathogenesis and progression of myocardial infarction (MI). Here we tried to investigate its potential mechanisms. The ratio of infarct size in heart issue was detected by TTC staining, and cardiac functions were calculated according to echocardiographic evaluation. Cell viability in cardiomyocytes was investigated by Cell Counting Kit-8 (CCK-8) and lactate dehydrogenase (LDH) release assays. Western blot was used to detect autophagy-related proteins including Beclin-1, LC3, p62, adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK)-mammalian target of rapamycin rabbit (mTOR)-p70 ribosomal protein S6 kinase (p70S6K) signaling-related proteins including AMPK, mTOR, p70S6K, and their phosphorylation formation. Rapamycin was used for the induction of autophagy. Cleaved caspase-3 and Bax were detected for analyzing apoptosis. TEM was used for the detection of autophagosomes. We found that CB2R deletion (CB2R KO) largely deteriorated the severity of MI and the cardiac function as well as cell viability of cardiomyocytes. Knocking out CB2R decreased the level of autophagy in heart issues from MI mice as well as cardiomyocytes under oxygen-glucose deprivation (OGD). Furthermore, CB2R dysfunction significantly attenuated the cardiac protective effects of rapamycin both in vivo and in vitro. Finally, we found that CB2R-mediated autophagy was induced by AMPK-mTOR-p70S6K signaling pathway. Our current study demonstrated for the first time that CB2R deletion led to a detrimental effect of MI through the dysfunction of AMPK-mTOR-p70S6K signaling pathway, which might provide a novel insight in the treatment of MI.

scholarly journals MiR-103a-3p antagonism attenuates pneumonia via inactivating the PI3K/AKT/NF-κB signaling pathway by targeting PTEN

2020 ◽  
Author(s):  
Lin Xu ◽  
Qingying Song ◽  
Zhanghong Ouyang ◽  
Xiangyan Zhang ◽  
Cheng Zhang
Keyword(s):  
Cell Viability ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Expression Levels ◽  
Tensin Homolog ◽  
Factor Α

Abstract Pneumonia accounts for approximately 15% mortalities in adolescents worldwide. MicroRNAs (miRNAs) regulate numerous diseases including pneumonia. miRNA and mRNA expression levels were detected by real time polymerase chain reaction (RT-qPCR). Protein expression levels were determined by enzyme-linked immunosorbent assay (ELISA) and western blot. The interaction between phosphatase and tensin homolog on chromosome ten (PTEN) and miR-103a-3p was explored by dual luciferase reporter assay. Cell viability and cell apoptosis were detected by cell Counting Kit-8 (CCK-8) and flow cytometry. Herein, we discovered that PTEN was decreased and miR-103a-3p was overexpressed in Ana-1 cells of in vitro pneumonia model. miR-103a-3p downregulated the expression levels of PTEN. AntagomiR-103a-3p reversed the increased cell apoptosis and decreased cell viability and inflammatory cytokine expression levels (tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6) induced by LPS in Ana-1 cells by PTEN. AntagomiR-103a-3p inhibited the activation of PTEN/PI3K/AKT/NF-κB signaling pathway induced by LPS in Ana-1 cells. Taken together, our findings exhibited that miR-103a-3p attenuated LPS induced pneumonia by blocking the activation of PTEN/PI3K/AKT/NF-κB signaling pathway and the following cell apoptosis as well as release of proinflammatory cytokines, suggesting that miR-103a-3p might serve as a novel therapeutic target for the treatment of pneumonia.

scholarly journals Cordycepin augments the chemosensitivity of osteosarcoma to cisplatin by activating AMPK and suppressing the AKT signaling pathway

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Hong-Bo Li ◽  
Jun-Kai Chen ◽  
Ze-Xin Su ◽  
Qing-Lin Jin ◽  
Li-Wen Deng ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Mtor Signaling ◽  
Osteosarcoma Cell ◽  
Mtor Signaling Pathway ◽  
Osteosarcoma Cells ◽  
Related Proteins

Abstract Background Osteosarcoma is the most common primary bone tumor in children and adolescents. However, some patients with osteosarcoma develop resistance to chemotherapy, leading to a poor clinical prognosis. Hence, effective therapeutic agents that can improve the response to chemotherapy drugs to improve the prognosis of patients with osteosarcoma are urgently needed. Cordycepin has recently emerged as a promising antitumor drug candidate. This study aims to explore the effect of cordycepin in suppressing osteosarcoma in vivo and in vitro and the synergistic effect of cordycepin combined with cisplatin and to demonstrate the underlying molecular mechanism. Methods CCK-8 assay was performed to investigate the inhibition effect of cordycepin combined with cisplatin in osteosarcoma cell lines. The colony formation and invasion abilities were measured by colony formation assay and Transwell assay. Osteosarcoma cells apoptosis was detected by flow cytometry. Western blot analysis were used to detect the expression of cell apoptosis-related proteins and AMPK and AKT/mTOR signaling pathway-related proteins. Finally, we performed the in vivo animal model to further explore whether cordycepin and cisplatin exert synergistic antitumor effects. Results Notably, we found that treatment with cordycepin inhibited cell proliferation, invasion, and induced apoptosis in osteosarcoma cells in vitro and in vivo. Moreover, the combination of cordycepin and cisplatin led to marked inhibition of osteosarcoma cell proliferation and invasion and promoted osteosarcoma cell apoptosis in vitro and in vivo. Mechanistically, we demonstrated that cordycepin enhanced the sensitivity of osteosarcoma cells to cisplatin by activating AMPK and inhibiting the AKT/mTOR signaling pathway. Conclusions In brief, this study provides comprehensive evidence that cordycepin inhibits osteosarcoma cell growth and invasion and induces osteosarcoma cell apoptosis by activating AMPK and inhibiting the AKT/mTOR signaling pathway and enhances the sensitivity of osteosarcoma cells to cisplatin, suggesting that cordycepin is a promising treatment for osteosarcoma.

Discovery of N-Phenyl-4-(1H-pyrrol-3-yl)pyrimidin-2-amine Derivatives as Potent Mnk2 Inhibitors: Design, Synthesis, SAR Analysis, and Evaluation of in vitro Anti-leukaemic Activity

2019 ◽  
Vol 15 (6) ◽  
pp. 602-623 ◽  
Author(s):  
Ahmed M. Abdelaziz ◽  
Sarah Diab ◽  
Saiful Islam ◽  
Sunita K.C. Basnet ◽  
Benjamin Noll ◽  
...  
Keyword(s):  
Proliferative Activity ◽  
Myeloid Cell ◽  
Structural Diversity ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cell Leukaemia ◽  
Aberrant Expression ◽  
Design Synthesis ◽  
Induced Apoptosis

Background:Aberrant expression of eukaryotic translation initiation factor 4E (eIF4E) is common in many types of cancer including acute myeloid leukaemia (AML). Phosphorylation of eIF4E by MAPK-interacting kinases (Mnks) is essential for the eIF4E-mediated oncogenic activity. As such, the pharmacological inhibition of Mnks can be an effective strategy for the treatment of cancer.Methods:A series of N-phenyl-4-(1H-pyrrol-3-yl)pyrimidin-2-amine derivatives was designed and synthesised. The Mnk inhibitory activity of these derivatives as well as their anti-proliferative activity against MV4-11 AML cells was determined.Results:These compounds were identified as potent Mnk2 inhibitors. Most of them demonstrated potent anti-proliferative activity against MV4-11 AML cells. The cellular mechanistic studies of the representative inhibitors revealed that they reduced the level of phosphorylated eIF4E and induced apoptosis by down-regulating the anti-apoptotic protein myeloid cell leukaemia 1 (Mcl-1) and by cleaving poly(ADP-ribose)polymerase (PARP). The lead compound 7k possessed desirable pharmacokinetic properties and oral bioavailability.Conclusion:This work proposes that exploration of the structural diversity in the context of Nphenyl- 4-(1H-pyrrol-3-yl)pyrimidin-2-amine would offer potent and selective Mnk inhibitors.

scholarly journals MicroRNA-144 Suppresses Prostate Cancer Growth and Metastasis by Targeting EZH2

2021 ◽  
Vol 20 ◽  
pp. 153303382198981
Author(s):  
Xin-bo Sun ◽  
Yong-wei Chen ◽  
Qi-sheng Yao ◽  
Xu-hua Chen ◽  
Min He ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Viability ◽  
Cell Apoptosis ◽  
Epithelial Mesenchymal Transition ◽  
Luciferase Reporter ◽  
Mesenchymal Transition ◽  
Proliferation And Apoptosis ◽  
High Incidence ◽  
Inhibit Cell Proliferation

Background: Prostate cancer is a common malignant tumor with a high incidence. MicroRNAs (miRNAs) have been shown to be important post-transcriptional regulators during tumorigenesis. This study aimed to explore the effect of miR-144 on PCa proliferation and apoptosis. Material and Methods: The expression of miR-144 and EZH2 were examined in clinical PCa tissues. PCa cell line LNCAP and DU-145 was employed and transfected with miR-144 mimics or inhibitors. The correlation between miR-144 and EZH2 was verified by luciferase reporter assay. Cell viability, apoptosis and migratory capacity were detected by CCK-8, flow cytometry assay and wound healing assay. The protein level of EZH2, E-Cadherin, N-Cadherin and vimentin were analyzed by western blotting. Results: miR-144 was found to be negatively correlated to the expression of EZH2 in PCa tissues. Further studies identified EZH2 as a direct target of miR-144. Moreover, overexpression of miR-144 downregulated expression of EZH2, reduced cell viability and promoted cell apoptosis, while knockdown of miR-144 led to an inverse result. miR-144 also suppressed epithelial-mesenchymal transition level of PCa cells. Conclusion: Our study indicated that miR-144 negatively regulate the expression of EZH2 in clinical specimens and in vitro. miR-144 can inhibit cell proliferation and induce cell apoptosis in PCa cells. Therefore, miR-144 has the potential to be used as a biomarker for predicting the progression of PCa.

scholarly journals Circadian clock control of eIF2α phosphorylation is necessary for rhythmic translation initiation

2020 ◽  
Vol 117 (20) ◽  
pp. 10935-10945 ◽  
Author(s):  
Shanta Karki ◽  
Kathrina Castillo ◽  
Zhaolan Ding ◽  
Olivia Kerr ◽  
Teresa M. Lamb ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Translation Initiation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Nutrient Metabolism ◽  
Eif2α Phosphorylation ◽  
Eif2α Kinase ◽  
The Impact

The circadian clock in eukaryotes controls transcriptional and posttranscriptional events, including regulation of the levels and phosphorylation state of translation factors. However, the mechanisms underlying clock control of translation initiation, and the impact of this potential regulation on rhythmic protein synthesis, were not known. We show that inhibitory phosphorylation of eIF2α (P-eIF2α), a conserved translation initiation factor, is clock controlled in Neurospora crassa, peaking during the subjective day. Cycling P-eIF2α levels required rhythmic activation of the eIF2α kinase CPC-3 (the homolog of yeast and mammalian GCN2), and rhythmic activation of CPC-3 was abolished under conditions in which the levels of charged tRNAs were altered. Clock-controlled accumulation of P-eIF2α led to reduced translation during the day in vitro and was necessary for the rhythmic synthesis of select proteins in vivo. Finally, loss of rhythmic P-eIF2α levels led to reduced linear growth rates, supporting the idea that partitioning translation to specific times of day provides a growth advantage to the organism. Together, these results reveal a fundamental mechanism by which the clock regulates rhythmic protein production, and provide key insights into how rhythmic translation, cellular energy, stress, and nutrient metabolism are linked through the levels of charged versus uncharged tRNAs.

scholarly journals Dual modulation of Ras-Mnk and PI3K-AKT-mTOR pathways: A Novel c-FLIP inhibitory mechanism of 3-AWA mediated translational attenuation through dephosphorylation of eIF4E

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Reyaz ur Rasool ◽  
Bilal Rah ◽  
Hina Amin ◽  
Debasis Nayak ◽  
Souneek Chakraborty ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Cell Cycle Progression ◽  
De Novo ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Apoptosis Resistance ◽  
Nih3t3 Cells ◽  
Eukaryotic Translation ◽  
Underlying Mechanisms

Abstract The eukaryotic translation initiation factor 4E (eIF4E) is considered as a key survival protein involved in cell cycle progression, transformation and apoptosis resistance. Herein, we demonstrate that medicinal plant derivative 3-AWA (from Withaferin A) suppressed the proliferation and metastasis of CaP cells through abrogation of eIF4E activation and expression via c-FLIP dependent mechanism. This translational attenuation prevents the de novo synthesis of major players of metastatic cascades viz. c-FLIP, c-Myc and cyclin D1. Moreover, the suppression of c-FLIP due to inhibition of translation initiation complex by 3-AWA enhanced FAS trafficking, BID and caspase 8 cleavage. Further ectopically restored c-Myc and GFP-HRas mediated activation of eIF4E was reduced by 3-AWA in transformed NIH3T3 cells. Detailed underlying mechanisms revealed that 3-AWA inhibited Ras-Mnk and PI3-AKT-mTOR, two major pathways through which eIF4E converges upon eIF4F hub. In addition to in vitro studies, we confirmed that 3-AWA efficiently suppressed tumor growth and metastasis in different mouse models. Given that 3-AWA inhibits c-FLIP through abrogation of translation initiation by co-targeting mTOR and Mnk-eIF4E, it (3-AWA) can be exploited as a lead pharmacophore for promising anti-cancer therapeutic development.

scholarly journals Hsp90 Binds and Regulates the Ligand-Inducible α Subunit of Eukaryotic Translation Initiation Factor Kinase Gcn2

1999 ◽  
Vol 19 (12) ◽  
pp. 8422-8432 ◽  
Author(s):  
Olivier Donzé ◽  
Didier Picard
Keyword(s):  
Amino Acid ◽  
Constitutive Expression ◽  
Amino Acid Starvation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Restrictive Temperature ◽  
Α Subunit ◽  
Macbecin I

ABSTRACT The protein kinase Gcn2 stimulates translation of the yeast transcription factor Gcn4 upon amino acid starvation. Using genetic and biochemical approaches, we show that Gcn2 is regulated by the molecular chaperone Hsp90 in budding yeast Saccharomyces cerevisiae. Specifically, we found that (i) several Hsp90 mutant strains exhibit constitutive expression of a GCN4-lacZ reporter plasmid; (ii) Gcn2 and Hsp90 form a complex in vitro as well as in vivo; (iii) the specific inhibitors of Hsp90, geldanamycin and macbecin I, enhance the association of Gcn2 with Hsp90 and inhibit its kinase activity in vitro; (iv) in vivo, macbecin I strongly reduces the levels of Gcn2; (v) in a strain expressing the temperature-sensitive Hsp90 mutant G170D, both the accumulation and activity of Gcn2 are abolished at the restrictive temperature; and (vi) the Hsp90 cochaperones Cdc37, Sti1, and Sba1 are required for the response to amino acid starvation. Taken together, these data identify Gcn2 as a novel target for Hsp90, which plays a crucial role for the maturation and regulation of Gcn2.

Translation in Saccharomyces cerevisiae: initiation factor 4E-dependent cell-free system

1989 ◽  
Vol 9 (10) ◽  
pp. 4467-4472
Author(s):  
M Altmann ◽  
N Sonenberg ◽  
H Trachsel
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Point Mutations ◽  
Translation Initiation Factor ◽  
Apparent Temperature ◽  
Initiation Factor ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Free System ◽  
Gal1 Promoter

The gene encoding translation initiation factor 4E (eIF-4E) from Saccharomyces cerevisiae was randomly mutagenized in vitro. The mutagenized gene was reintroduced on a plasmid into S. cerevisiae cells having their only wild-type eIF-4E gene on a plasmid under the control of the regulatable GAL1 promoter. Transcription from the GAL1 promoter (and consequently the production of wild-type eIF-4E) was then shut off by plating these cells on glucose-containing medium. Under these conditions, the phenotype conferred upon the cells by the mutated eIF-4E gene became apparent. Temperature-sensitive S. cerevisiae strains were identified by replica plating. The properties of one strain, 4-2, were further analyzed. Strain 4-2 has two point mutations in the eIF-4E gene. Upon incubation at 37 degrees C, incorporation of [35S]methionine was reduced to 15% of the wild-type level. Cell-free translation systems derived from strain 4-2 were dependent on exogenous eIF-4E for efficient translation of certain mRNAs, and this dependence was enhanced by preincubation of the extract at 37 degrees C. Not all mRNAs tested required exogenous eIF-4E for translation.

scholarly journals Intermittent-Hypoxia-Induced Autophagy Activation Through the ER-Stress-Related PERK/eIF2α/ATF4 Pathway is a Protective Response to Pancreatic β-Cell Apoptosis

2018 ◽  
Vol 51 (6) ◽  
pp. 2955-2971 ◽  
Author(s):  
Shuling Song ◽  
Jin Tan ◽  
Yuyang Miao ◽  
Zuoming Sun ◽  
Qiang  Zhang
Keyword(s):  
Er Stress ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Intermittent Hypoxia ◽  
Autophagic Vacuole ◽  
Pancreatic Β Cell ◽  
Β Cell ◽  
Protective Response

Background/Aims: Intermittent hypoxia (IH) causes apoptosis in pancreatic β-cells, but the potential mechanisms remain unclear. Endoplasmic reticulum (ER) stress, autophagy, and apoptosis are interlocked in an extensive crosstalk. Thus, this study aimed to investigate the contributions of ER stress and autophagy to IH-induced pancreatic β-cell apoptosis. Methods: We established animal and cell models of IH, and then inhibited autophagy and ER stress by pharmacology and small interfering RNA (siRNA) in INS-1 cells and rats. The levels of biomarkers for autophagy, ER stress, and apoptosis were evaluated by immunoblotting and immunofluorescence. The number of autophagic vacuoles was observed by transmission electron microscopy. Results: IH induced autophagy activation both in vivo and in vitro, as evidenced by increased autophagic vacuole formation and LC3 turnover, and decreased SQSTM1 level. The levels of ER-stress-related proteins, including GRP78, CHOP, caspase 12, phosphorylated (p)-protein kinase RNA-like ER kinase (PERK), p-eIF2α, and activating transcription factor 4 (ATF4) were increased under IH conditions. Inhibition of ER stress with tauroursodeoxycholic acid or 4-phenylbutyrate partially blocked IH-induced autophagy in INS-1 cells. Furthermore, inhibition of PERK with GSK2606414 or siRNA blocked the ERstress-related PERK/eIF2α/ATF4 signaling pathway and inhibited autophagy induced by IH, which indicates that IH-induced autophagy activation is dependent on this signaling pathway. Promoting autophagy with rapamycin alleviated IH-induced apoptosis, whereas inhibition of autophagy with chloroquine or autophagy-related gene (Atg5 and Atg7) siRNA aggravated pancreatic β-cell apoptosis caused by IH. Conclusion: IH induces autophagy activation through the ER-stress-related PERK/eIF2α/ATF4 signaling pathway, which is a protective response to pancreatic β-cell apoptosis caused by IH.

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