• International Journal of Oral Biology
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• 제45권4호
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• pp.190-196
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• 2020

Several factors, including genetic and environmental insults, impede protein folding and secretion in the endoplasmic reticulum (ER). Accumulation of unfolded or mis-folded protein in the ER manifests as ER stress. To cope with this morbid condition of the ER, recent data has suggested that the intracellular event of an unfolded protein response plays a critical role in managing the secretory load and maintaining proteostasis in the ER. Tauroursodeoxycholic acid (TUDCA) is a chemical chaperone and hydrophilic bile acid that is known to inhibit apoptosis by attenuating ER stress. Numerous studies have revealed that TUDCA affects hepatic diseases, obesity, and inflammatory illnesses. Recently, molecular regulation of ER stress in tooth development, especially during the secretory stage, has been studied. Therefore, in this study, we examined the developmental role of ER stress regulation in tooth morphogenesis using in vitro organ cultivation methods with a chemical chaperone treatment, TUDCA. Altered cellular events including proliferation, apoptosis, and dentinogenesis were examined using immunostaining and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. In addition, altered localization patterns of the formation of hard tissue matrices related to molecules, including amelogenin and nestin, were examined to assess their morphological changes. Based on our findings, modulating the role of the chemical chaperone TUDCA in tooth morphogenesis, especially through the modulation of cellular proliferation and apoptosis, could be applied as a supporting data for tooth regeneration for future studies.

• 대한의생명과학회지
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• 제16권1호
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• pp.65-67
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• 2010

The endoplasmic reticulum (ER) is a multifunctional intercellular organelle in which several posttranslational modification steps occurred such as protein folding, lipid biosynthesis, calcium storage and release. Perturbations that disrupt ER homeostasis lead to the misfolding of proteins in the ER lumen and up-regulation of ER signaling pathway called the unfolded protein response (UPR). Here, we have demonstrated that ageing changes the expression of ER chaperone and associated ER membrane kinases of IRE1, ATF6 and PERK.

• BMB Reports
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• 제53권11호
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• pp.576-581
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• 2020

Dimethylation of the histone H3 protein at lysine residue 9 (H3K9) is mediated by euchromatin histone methyltransferase II (EHMT2) and results in transcriptional repression of target genes. Recently, chemical inhibition of EHMT2 was shown to induce various physiological outcomes, including endoplasmic reticulum stress-associated genes transcription in cancer cells. To identify genes that are transcriptionally repressed by EHMT2 during apoptosis, and cell stress responses, we screened genes that are upregulated by BIX-01294, a chemical inhibitor of EHMT2. RNA sequencing analyses revealed 77 genes that were upregulated by BIX-01294 in all four hepatic cell carcinoma (HCC) cell lines. These included genes that have been implicated in apoptosis, the unfolded protein response (UPR), and others. Among these genes, the one encoding the stress-response protein Ras-related GTPase C (RRAGC) was upregulated in all BIX-01294-treated HCC cell lines. We confirmed the regulatory roles of EHMT2 in RRAGC expression in HCC cell lines using proteomic analyses, chromatin immune precipitation (ChIP) assay, and small guide RNA-mediated loss-of-function experiments. Upregulation of RRAGC was limited by the reactive oxygen species (ROS) scavenger N-acetyl cysteine (NAC), suggesting that ROS are involved in EHMT2-mediated transcriptional regulation of stress-response genes in HCC cells. Finally, combined treatment of cells with BIX-01294 and 5-Aza-cytidine induced greater upregulation of RRAGC protein expression. These findings suggest that EHMT2 suppresses expression of the RRAGC gene in a ROS-dependent manner and imply that EHMT2 is a key regulator of stress-responsive gene expression in liver cancer cells.

• Journal of Microbiology and Biotechnology
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• 제23권12호
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• pp.1785-1790
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• 2013

The synthetic machinery of ATF4 (activating transcription factor 4) is activated in response to various stress conditions involved in nutrient restriction, endoplasmic reticulum homeostasis, and oxidation. Stress-induced inhibition of proteasome activity triggers the unfolded protein response and endoplasmic reticulum stress, where ATF4 is crucial for consequent biological events. In the current study, we showed that the $NAD^+$-dependent deacetylase, SIRT1, suppresses ATF4 synthesis during proteasome inhibition. SIRT1 depletion via transfection of specific siRNA into HeLa cells resulted in a significant increase in ATF4 protein, which was observed specifically in the presence of the proteasome inhibitor MG132. Consistent with SIRT1 depletion data, transient transfection of cells with SIRT1-overexpressing plasmid induced a decrease in the ATF4 protein level in the presence of MG132. Interestingly, however, ATF4 mRNA was not affected by SIRT1, even in the presence of MG132, indicating that SIRT1-induced suppression of ATF4 synthesis occurs under post-transcriptional control. Accordingly, we propose that SIRT1 serves as a negative regulator of ATF4 protein synthesis at the post-transcriptional level, which is observed during stress conditions, such as proteasome inhibition.

• 대한의생명과학회지
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• 제23권4호
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• pp.416-420
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• 2017

Endoplasmic reticulum (ER) stress induces unfolded protein response (UPR) via inositol-requiring enzyme 1 (IRE1) activation, which sends a molecular signal for X box-binding protein 1 (XBP1) mRNA splicing in the cytosol. IRE1 endoribonuclease activity induces cleavage of XBP1 mRNA. The XBP1 mRNA is then ligated by an uncharacterized RNA ligase and translated to produce spliced XBP1 by 23 nt removed in which contains the PstI restriction enzyme site. The splicing of XBP1 mRNA can be detected by semiquantitative RT-PCR, and then splicing of XBP1 is a useful tool to measure the genetic variability in ER stress. In this study, we have estimated IRE1-dependent splicing of XBP1 mRNA under conditions of various hypothermia. The results indicated that hypothermia regulated ER stress. This study demonstrated that hypothermia is closely related to ER stress and may be useful for early diagnosis of ER-associated disease.

• Molecules and Cells
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• 제26권4호
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• pp.323-328
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• 2008

Eukaryotic cells continuously integrate intrinsic and extrinsic signals to adapt to the environment. When exposed to stressful conditions, cells activate compartment-specific adaptive responses. If these are insufficient, apoptosis ensues as an organismal defense line. The mechanisms that sense stress and set the transition from adaptive to maladaptive responses, activating apoptotic programs, are the subject of intense studies, also for their potential impact in cancer and degenerative disorders. In the former case, one would aim at lowering the threshold, in the latter instead to increase it. Protein synthesis, consuming energy for anabolic processes as well as for byproducts disposal, can be a significant source of stress, particularly when difficult-to-fold proteins are produced. Recent work from our and other laboratories on the differentiation of antibody secreting cells, revealed a regulatory circuit that integrates protein synthesis, secretion and degradation (proteostasis), into cell lifespan determination. The apoptotic elimination - after an industrious, yet short lifetime - of terminal immune effectors is crucial to maintain immune homeostasis. Linking proteostasis to cell death, this paradigm might prove useful for biotechnological purposes, and the design of novel anti-cancer therapies.

• 생명과학회지
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• 제30권4호
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• pp.402-409
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• 2020

신경괴사증바이러스(NNV)는 25 nm의 작은 입자 크기에 RNA1 (3.4 kb, RdRp), RNA2 (1.4 kb, capsid protein) 두 가닥의 RNA를 유전정보를 가진다. NNV는 1980년대 말 처음 보고된 이후 전 세계적으로 120여종의 어류에 감염을 일으키며 심각한 피해를 일으키고 있는 바이러스이다. NNV 감염에 의한 피해를 최소화하고 효율적인 백신들을 개발하기 위해서는 무엇보다 NNV 감염에 따른 세포내 신호전달체계를 이해할 필요가 있다. NNV는 세포내 감염 이후 숙주가 가진 바이러스 복제에 필요한 요소들을 이용할 수 있도록 숙주세포의 cell cycle arrest 등의 기작을 이용하는 것으로 알려졌다. 반면에 숙주 세포는 NNV와 감염된 세포를 제어하기 위해 RIG-1-like receptor signaling pathway 등을 통해 NNV 감염을 인지한 다음 IFN signaling pathway를 통해 항바이러스 작용에 필요한 ISG들을 발현시킨다. 또한 감염된 세포들을 사멸시키기 위해 ER stress를 통한 unfolded protein response (UPR), mitochondria-mediated cell death 작용을 통해 감염된 세포의 apoptosis를 유발한다. NNV 감염 기작에 대한 세포신호전달연구는 아직 초기단계이며 검증해야 할 pathway들이 아직도 많이 남아있는 상황이다. 따라서 NNV 감염과 연관된 다양한 세포신호전달체계를 탐색하고 질병 특이적인 세포신호전달체계를 이해함으로써 신속하고 정확한 진단법 및 백신 개발에 많은 도움이 될 것으로 생각된다.

• Molecules and Cells
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• 제40권7호
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• pp.457-465
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• 2017

Streptozotocin (STZ)-induced murine models of type 1 diabetes have been used to examine ER stress during pancreatic ${\beta}$-cell apoptosis, as this ER stress plays important roles in the pathogenesis and development of the disease. However, the mechanisms linking type 1 diabetes to the ER stress-modulating anti-diabetic signaling pathway remain to be addressed, though it was recently established that ERK5 (Extracellular-signal-regulated kinase 5) contributes to the pathogeneses of diabetic complications. This study was undertaken to explore the mechanism whereby ERK5 inhibition instigates pancreatic ${\beta}$-cell apoptosis via an ER stress-dependent signaling pathway. STZ-induced diabetic WT and CHOP deficient mice were i.p. injected every 2 days for 6 days under BIX02189 (a specific ERK5 inhibitor) treatment in order to evaluate the role of ERK5. Hyperglycemia was exacerbated by co-treating C57BL/6J mice with STZ and BIX02189 as compared with mice administered with STZ alone. In addition, immunoblotting data revealed that ERK5 inhibition activated the unfolded protein response pathway accompanying apoptotic events, such as, PARP-1 and caspase-3 cleavage. Interestingly, ERK5 inhibition-induced exacerbation of pancreatic ${\beta}$-cell apoptosis was inhibited in CHOP deficient mice. Moreover, transduction of adenovirus encoding an active mutant form of $MEK5{\alpha}$, an upstream kinase of ERK5, inhibited STZ-induced unfolded protein responses and ${\beta}$-cell apoptosis. These results suggest that ERK5 protects against STZ-induced pancreatic ${\beta}$-cell apoptosis and hyperglycemia by interrupting the ER stress-mediated apoptotic pathway.

• BMB Reports
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• 제45권8호
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• pp.482-487
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• 2012

To identify the novel inhibitors of endoplasmic reticulum stress-induced cell death, we performed a high throughput assay with a chemical library containing a total of 3,280 bioactive small molecules. Cyclosporine A and bromocriptine were identified as potent inhibitors of thapsigargiin-induced cell death (cut-off at $4{\sigma}$ standard score). However, U74389G, the potent inhibitor of lipid peroxidation had lower activity in inhibiting cell death. The inhibition effect of cyclosporine A and bromocriptine was specific for only thapsigargin-induced cell death. The mechanism of inhibition by these compounds was identified as modification of the expression of glucose regulated protein-78 (GRP-78/Bip) and inhibition of phosphorylation of p38 mitogen activated protein kinase (MAPK). However, these compounds did not inhibit the same events triggered by tunicamycin, which was in agreement with the cell survival data. We suggest that the induction of protective unfolded protein response by these compounds confers resistance to cell death. In summary, we identified compounds that may provide insights on cell death mechanisms stimulated by ER stress.

• Toxicological Research
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• 제24권2호
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• pp.129-135
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• 2008

The results of recent studies indicate that high levels of free fatty acids(FFAs) and adipokines may be the main causes of non-alcoholic liver disease; however, the molecular mechanism that links FFAs to lipotoxicity remains unclear. In the present study, we treated HepG2 cells with FFA(either palmitate or oleate) to investigate the mechanisms involved in lipotoxicity in the liver cells. We also treated cells with palmitate in the presence of a chemical chaperone, 4-phenylbutyric acid(PBA), to confirm the involvement of ER stress in lipotoxicity. Palmitate significantly induced cytotoxicity in dose- and time-dependent manners. Apoptosis was also significantly induced by palmitate as measured by caspase-3 activity and DAPI staining. Palmitate led to increased expressions of the spliced form of X-box-protein(Xbp)-1 mRNA and C/EBP homologous transcription factor(CHOP) protein, suggesting activation of the unfolded-protein response. PBA co-incubation significantly attenuated apoptosis induced by palmitate. The above data demonstrate that high levels of palmitate induce apoptosis via the mediation of ER stress in the liver cells and that chemical chaperones act to modulate ER stress and accompanying apoptosis.