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Autophagy Oct 2014Transient cerebral ischemia leads to endoplasmic reticulum (ER) stress. However, the contributions of ER stress to cerebral ischemia are not clear. To address this...
Transient cerebral ischemia leads to endoplasmic reticulum (ER) stress. However, the contributions of ER stress to cerebral ischemia are not clear. To address this issue, the ER stress activators tunicamycin (TM) and thapsigargin (TG) were administered to transient middle cerebral artery occluded (tMCAO) mice and oxygen-glucose deprivation-reperfusion (OGD-Rep.)-treated neurons. Both TM and TG showed significant protection against ischemia-induced brain injury, as revealed by reduced brain infarct volume and increased glucose uptake rate in ischemic tissue. In OGD-Rep.-treated neurons, 4-PBA, the ER stress releasing mechanism, counteracted the neuronal protection of TM and TG, which also supports a protective role of ER stress in transient brain ischemia. Knocking down the ER stress sensor Eif2s1, which is further activated by TM and TG, reduced the OGD-Rep.-induced neuronal cell death. In addition, both TM and TG prevented PARK2 loss, promoted its recruitment to mitochondria, and activated mitophagy during reperfusion after ischemia. The neuroprotection of TM and TG was reversed by autophagy inhibition (3-methyladenine and Atg7 knockdown) as well as Park2 silencing. The neuroprotection was also diminished in Park2(+/-) mice. Moreover, Eif2s1 and downstream Atf4 silencing reduced PARK2 expression, impaired mitophagy induction, and counteracted the neuroprotection. Taken together, the present investigation demonstrates that the ER stress induced by TM and TG protects against the transient ischemic brain injury. The PARK2-mediated mitophagy may be underlying the protection of ER stress. These findings may provide a new strategy to rescue ischemic brains by inducing mitophagy through ER stress activation.
Topics: Animals; Apoptosis; Brain Ischemia; Endoplasmic Reticulum Stress; Glucose; Mice; Mitophagy; Neuroprotective Agents; Oxygen; Reperfusion Injury; Signal Transduction; Thapsigargin; Tunicamycin; Ubiquitin-Protein Ligases
PubMed: 25126734
DOI: 10.4161/auto.32136 -
Journal of Experimental & Clinical... Nov 2018Multidrug resistance remains a major obstacle to successful treatment for patients with gastric cancer (GC). Recently, glycosylation has been demonstrated to play a...
BACKGROUND
Multidrug resistance remains a major obstacle to successful treatment for patients with gastric cancer (GC). Recently, glycosylation has been demonstrated to play a vital role in the acquisition of multidrug resistance. As a potent inhibitor of glycosylation, tunicamycin (Tu) has shown marked antitumor activities in various cancers. In the present study, we attempted to determine the exact effect of Tu on the chemoresistance of GC.
METHODS
The cytotoxic effects of drugs on GC cells were evaluated by cell viability assays, and apoptosis was detected by flow cytometry. PCR, western blot analysis, immunofluorescence staining and canonical inhibitors were employed to identify the underlying mechanisms of the specific effects of Tu on multidrug-resistant (MDR) GC cells.
RESULTS
For the first time, we found that MDR GC cells were more sensitive to Tu-induced cell death than the parental cells and that the increased sensitivity might correlate with basal endoplasmic reticulum (ER) stress. In addition, Tu dramatically increased chemotherapy-induced apoptosis by evoking ER stress in GC cells, particularly MDR cells. Further study indicated that these effects were highly dependent on glycosylation inhibition by Tu, rather than its role as a canonical ER stress inducer. Besides, autophagy was markedly triggered by Tu, and blocking autophagy enhanced the combined effects of Tu and chemotherapy on MDR GC cells.
CONCLUSIONS
Our results suggest that tumor-targeted glycosylation inhibition may be a feasible strategy to reverse chemoresistance in GC patients.
Topics: Anti-Bacterial Agents; Apoptosis; Cell Line, Tumor; Drug Resistance, Neoplasm; Endoplasmic Reticulum Stress; Glycosylation; Humans; Stomach Neoplasms; Tunicamycin
PubMed: 30413206
DOI: 10.1186/s13046-018-0935-8 -
Molecules and Cells Jun 2023N-glycosylation, a common post-translational modification, is widely acknowledged to have a significant effect on protein stability and folding. N-glycosylation is a... (Review)
Review
N-glycosylation, a common post-translational modification, is widely acknowledged to have a significant effect on protein stability and folding. N-glycosylation is a complex process that occurs in the endoplasmic reticulum (ER) and requires the participation of multiple enzymes. GlcNAc-1-P-transferase (GPT) is essential for initiating N-glycosylation in the ER. Tunicamycin is a natural product that inhibits N-glycosylation and produces ER stress, and thus it is utilized in research. The molecular mechanism by which GPT triggers N-glycosylation is discussed in this review based on the GPT structure. Based on the structure of the GPT-tunicamycin complex, we also discuss how tunicamycin reduces GPT activity, which prevents N-glycosylation. This review will be highly useful for understanding the role of GPT in the N-glycosylation of proteins, as well as presents a potential for considering tunicamycin as an antibiotic treatment.
Topics: Glycosylation; Tunicamycin; Protein Processing, Post-Translational; Endoplasmic Reticulum Stress; Endoplasmic Reticulum
PubMed: 37190766
DOI: 10.14348/molcells.2023.0001 -
International Journal of Molecular... Mar 2023Endothelial dysfunction (ED) is the initiation of atherosclerosis (AS). Our previous studies have found that cholesterol metabolism and the Wnt/β-catenin pathway can...
Endothelial dysfunction (ED) is the initiation of atherosclerosis (AS). Our previous studies have found that cholesterol metabolism and the Wnt/β-catenin pathway can affect endoplasmic reticulum stress (ER stress), which ultimately leads to ED. However, the effects of cholesterol efflux on ED, which are caused by oxidative stress and the correlation among ER stress, Wnt/β-catenin pathway, and cholesterol efflux, are not clear during ED. To uncover them, the expressions of liver X receptors (LXRα and LXRβ) and ATP-binding cassette protein A1 (ABCA1) and G1 (ABCG1) in HUVECs (human umbilical vein endothelial cells) were measured under oxidative stress. Moreover, HUVECs were treated with LXR-623 (LXR agonist), cholesterol, tunicamycin, and salinomycin alone or together. The results indicated that oxidative stress-induced ED could deregulate the expressions of LXRα and LXRβ and trigger the ER stress and Wnt/β-catenin pathway, resulting thereafter in the accumulation of cholesterol. Furthermore, similar results were shown after treatment with cholesterol; however, the activation of liver X receptor (LXR) could reverse these changes. Furthermore, other results demonstrated that tunicamycin-induced ER stress could stimulate the accumulation of cholesterol and the Wnt/β-catenin pathway, further leading to ED. Inversely, salinomycin could reverse the above effects by deregulating the Wnt/β-catenin pathway. Collectively, our results showed that cholesterol efflux is partly responsible for the oxidative stress-induced ED; in addition, ER stress, the Wnt/β-catenin pathway, and cholesterol metabolism can interact with each other to promote ED.
Topics: Humans; beta Catenin; Tunicamycin; Liver X Receptors; Cholesterol; Human Umbilical Vein Endothelial Cells; Oxidative Stress; ATP Binding Cassette Transporter 1
PubMed: 36983012
DOI: 10.3390/ijms24065939 -
Biomolecules Mar 2022Breast cancer is an ancient disease recognized first by the Egyptians as early as 1600 BC. The first cancer-causing gene in a chicken tumor virus was found in 1970. The... (Review)
Review
Breast cancer is an ancient disease recognized first by the Egyptians as early as 1600 BC. The first cancer-causing gene in a chicken tumor virus was found in 1970. The United States signed the National Cancer Act in 1971, authorizing federal funding for cancer research. Irrespective of multi-disciplinary approaches, diverting a great deal of public and private resources, breast cancer remains at the forefront of human diseases, affecting as many as one in eight women during their lifetime. Because of overarching challenges and changes in the breast cancer landscape, five-year disease-free survival is no longer considered adequate. The absence of a cure, and the presence of drug resistance, severe side effects, and destruction of the patient's quality of life, as well as the fact that therapy is often expensive, making it unaffordable to many, have created anxiety among patients, families, and friends. One of the reasons for the failure of cancer therapeutics is that the approaches do not consider cancer holistically. Characteristically, all breast cancer cells and their microenvironmental capillary endothelial cells express asparagine-linked (-linked) glycoproteins with diverse structures. We tested a small biological molecule, Tunicamycin, that blocks a specific step of the protein -glycosylation pathway in the endoplasmic reticulum (ER), i.e., the catalytic activity of -acetylglusosaminyl 1-phosphate transferase (GPT). The outcome was overwhelmingly exciting. Tunicamycin quantitatively inhibits angiogenesis in vitro and in vivo, and inhibits the breast tumor progression of multiple subtypes in pre-clinical mouse models with "zero" toxicity. Mechanistic details support ER stress-induced unfolded protein response () signaling as the cause for the apoptotic death of both cancer and the microvascular endothelial cells. Additionally, it interferes with Wnt signaling. We therefore conclude that Tunicamycin can be expected to supersede the current therapeutics to become a glycotherapy for treating breast cancer of all subtypes.
Topics: Animals; Breast Neoplasms; Endoplasmic Reticulum Stress; Endothelial Cells; Female; Humans; Mice; Quality of Life; Tunicamycin; Unfolded Protein Response
PubMed: 35454076
DOI: 10.3390/biom12040487 -
Cell Death & Disease Aug 2023Unfolded protein response (UPR) maintains the endoplasmic reticulum (ER) homeostasis, survival, and physiological function of mammalian cells. However, how cells adapt...
Unfolded protein response (UPR) maintains the endoplasmic reticulum (ER) homeostasis, survival, and physiological function of mammalian cells. However, how cells adapt to ER stress under physiological or disease settings remains largely unclear. Here by a genome-wide CRISPR screen, we identified that RBBP8, an endonuclease involved in DNA damage repair, is required for ATF4 activation under ER stress in vitro. RNA-seq analysis suggested that RBBP8 deletion led to impaired cell cycle progression, retarded proliferation, attenuated ATF4 activation, and reduced global protein synthesis under ER stress. Mouse tissue analysis revealed that RBBP8 was highly expressed in the liver, and its expression is responsive to ER stress by tunicamycin intraperitoneal injection. Hepatocytes with RBBP8 inhibition by adenovirus-mediated shRNA were resistant to tunicamycin (Tm)-induced liver damage, cell death, and ER stress response. To study the pathological role of RBBP8 in regulating ATF4 activity, we illustrated that both RBBP8 and ATF4 were highly expressed in liver cancer tissues compared with healthy controls and highly expressed in Ki67-positive proliferating cells within the tumors. Interestingly, overexpression of RBBP8 in vitro promoted ATF4 activation under ER stress, and RBBP8 expression showed a positive correlation with ATF4 expression in liver cancer tissues by co-immunostaining. Our findings provide new insights into the mechanism of how cells adapt to ER stress through the crosstalk between the nucleus and ER and how tumor cells survive under chemotherapy or other anticancer treatments, which suggests potential therapeutic strategies against liver disease by targeting DNA damage repair, UPR or protein synthesis.
Topics: Animals; Mice; Clustered Regularly Interspaced Short Palindromic Repeats; Tunicamycin; Unfolded Protein Response; Liver Neoplasms; Mammals
PubMed: 37591836
DOI: 10.1038/s41419-023-06046-x -
Microbial Pathogenesis Dec 2020SARS-CoV-2 remains a medical and economic challenge, due to the lack of a suitable drug or vaccine. The glycans in some proteins play a pivotal role in protein folding,... (Review)
Review
SARS-CoV-2 remains a medical and economic challenge, due to the lack of a suitable drug or vaccine. The glycans in some proteins play a pivotal role in protein folding, oligomerization, quality control, sorting, and transport so the hindering of N-linked glycosylation of glycoproteins will prevent assembly of the virion. Tunicamycin an anticancer drug inhibit the N- linked glycans. Our study aimed to find out the mechanism action of tunicamycin on the viral glycoproteins. The growth of coronavirus in the presence inhibitor tunicamycin resulted in the production of spikeless, non-infectious virions which were devoid of S protein. We concluded that tunicamycin inhibits E2, S, and M glycoproteins of coronaviruses. Tunicamycin is also diminished glycosylation of PTMs such as HE, and 8 ab of SARS-CoV. Finally, we recommend using this drug to treat the SARS-CoV-2.
Topics: Animals; COVID-19; Glycosylation; Humans; SARS-CoV-2; Spike Glycoprotein, Coronavirus; Tunicamycin; Viral Matrix Proteins; COVID-19 Drug Treatment
PubMed: 33091582
DOI: 10.1016/j.micpath.2020.104586 -
Investigative Ophthalmology & Visual... Nov 2023Endoplasmic reticulum (ER) and mitochondrial stress are independently associated with corneal endothelial cell (CEnC) loss in many corneal diseases, including Fuchs'...
PURPOSE
Endoplasmic reticulum (ER) and mitochondrial stress are independently associated with corneal endothelial cell (CEnC) loss in many corneal diseases, including Fuchs' endothelial corneal dystrophy (FECD). However, the role of ER stress in mitochondrial dysfunction contributing to CEnC apoptosis is unknown. The purpose of this study is to explore the crosstalk between ER and mitochondrial stress in CEnC.
METHODS
Human corneal endothelial cell line (HCEnC-21T) and human corneal endothelial tissues were treated with ER stressor tunicamycin. ER stress-reducing chemical 4-phenyl butyric acid (4-PBA) was used in HCEnC-21T after tunicamycin. Fuchs' corneal endothelial cell line (F35T) was used to determine differential activation of ER stress with respect to HCEnC-21T at the baseline. ER stress, mitochondrial-mediated intrinsic apoptotic, mitochondrial fission, and fusion proteins were determined using immunoblotting and immunohistochemistry. Mitochondrial bioenergetics were assessed by mitochondrial membrane potential (MMP) loss and ATP production at 48 hours after tunicamycin. Mitochondria dynamics (shape, area, perimeter) were also analyzed at 24 hours using transmission electron microscopy.
RESULTS
Treatment of HCEnC-21T cell line with tunicamycin activated three ER stress pathways (PERK-eIF2α-CHOP, IRE1α-XBP1, and ATF6), reduced cell viability, upregulated mitochondrial-mediated intrinsic apoptotic molecules (cleaved caspase 9, caspase 3, PARP, Bax, cytochrome C), downregulated anti-apoptotic Bcl-2 protein, initiated mitochondrial dysfunction by loss of MMP and lowering of ATP production, and caused mitochondrial swelling and fragmentation with increased expression of mitochondrial fission proteins (Fis1 and p-Drp1). Fuchs' CEnC (F35T) cell line also showed activation of the ER stress-related proteins (p-eIF2α, GRP78, CHOP, XBP1) compared to HCEnC-21T at the baseline. The 4-PBA ameliorated cell loss and reduced cleaved caspase 3 and 9, thereby rescuing tunicamycin-induced cell death but not mitochondrial bioenergetics in HCEnC-21T cell line.
CONCLUSIONS
Tunicamycin-induced ER stress disrupts mitochondrial bioenegetics, dynamics and contributes to the loss of CEnC viability. This novel study highlights the importance of ER-mitochondria crosstalk and its contribution to CEnCs apoptosis, seen in many corneal diseases, including FECD.
Topics: Humans; Caspase 3; Endoribonucleases; Tunicamycin; Protein Serine-Threonine Kinases; Apoptosis; Endoplasmic Reticulum Stress; Corneal Diseases; Fuchs' Endothelial Dystrophy; Butyric Acid; Energy Metabolism; Endothelial Cells; Adenosine Triphosphate
PubMed: 37962528
DOI: 10.1167/iovs.64.14.18 -
MBio Aug 2021How cells exposed to one stress are later able to better survive other types of stress is not well understood. In eukaryotic organisms, physiological and pathological...
How cells exposed to one stress are later able to better survive other types of stress is not well understood. In eukaryotic organisms, physiological and pathological stresses can disturb endoplasmic reticulum (ER) function, resulting in "ER stress." Here, we found that exposure to tunicamycin, an inducer of ER stress, resulted in the acquisition of a specific aneuploidy, chromosome 2 trisomy (Chr2x3), in Candida albicans. Importantly, the resulting aneuploidy also conferred cross-tolerance to caspofungin, a first-line echinocandin antifungal, as well as to hydroxyurea, a common chemotherapeutic agent. Exposure to a range of tunicamycin concentrations induced similar ER stress responses. Extra copies of one Chr2 gene, , affected both tunicamycin and caspofungin tolerance, while at least 3 genes on chromosome 2 (, , and ) affected only tunicamycin and not caspofungin responses. Other Chr2 genes ( and ) affected hydroxyurea tolerance but neither tunicamycin nor caspofungin tolerance. Deletion of components of the protein kinase C (PKC) or calcineurin pathways affected tolerance to both tunicamycin and caspofungin, supporting the idea that the ER stress response and echinocandin tolerance are regulated by overlapping stress response pathways. Thus, antifungal drug tolerance can arise rapidly via ER stress-induced aneuploidy. Candida albicans is a prevalent human fungal commensal and also a pathogen that causes life-threatening systemic infections. Treatment failures are frequent because few therapeutic antifungal drug classes are available and because drug resistance and tolerance limit drug efficacy. We found that C. albicans rapidly overcomes the cellular stress induced by the drug tunicamycin by duplicating chromosome 2. Also, chromosome 2 duplication confers tolerance not only to tunicamycin but also to the following two unrelated drugs: caspofungin, an antifungal drug, and hydroxyurea, a chemotherapeutic. Cross tolerance to the three drugs involves different sets of genes, although some genetic pathways affect the tolerance to two of these three drugs. This work highlights a serious concern, namely, that changes in whole chromosome copy number can occur in response to one type of stress, and yet, they may facilitate the emergence of tolerance to multiple drugs, including the few antifungal drug classes available to treat infections.
Topics: Aneuploidy; Antifungal Agents; Candida albicans; Drug Resistance, Fungal; Stress, Physiological; Tunicamycin
PubMed: 34465026
DOI: 10.1128/mBio.02272-21 -
Journal of Healthcare Engineering 2021Triple negative breast cancer (TNBC) has significantly threatened human health. Many aspects of TNBC are closely related to Wnt/-catenin pathway, and cell apoptosis...
Triple negative breast cancer (TNBC) has significantly threatened human health. Many aspects of TNBC are closely related to Wnt/-catenin pathway, and cell apoptosis induced by endoplasmic reticulum stress (ER stress) in TNBC may act as a potential target of non-chemotherapy treatment. However, how ER stress interacts with this pathway in TNBC has not yet been understood. Here, the tunicamycin and LiCl have been applied to MDA-MB-231. The related proteins' expression was measured by western blotting. Moreover, acridine orange/ethidium bromide (AO/EB) staining was applied to test the apoptosis degree of the cells, and cell viability was tested by MTT experiment. Then, we found the ER stress and apoptosis degree of MDA-MB-231 were induced after treatment with tunicamycin. Besides, tunicamycin dose dependently inhibited both Wnt/-catenin pathway and cells viability. Licl, an activator of Wnt/-catenin signaling pathway, could significantly inhibit cell apoptosis. In conclusion, our study found that the activation of ER stress could promote the MDA-MB-231 apoptosis by repressing Wnt/-catenin pathway, which provides some promising prospects and basic mechanism to the further research.
Topics: Apoptosis; Cell Line, Tumor; Cell Proliferation; Endoplasmic Reticulum Stress; Humans; Triple Negative Breast Neoplasms; Tunicamycin; Wnt Signaling Pathway
PubMed: 34336161
DOI: 10.1155/2021/6394514