• Journal of Life Science
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• v.9 no.1
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• pp.76-83
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• 1999

Mammalial expression vectors containing GRP94, BiP, ERp72, and PDI, were introduced into FRTL5 cells. Transfected cells were selected by neomycin resistance for exogenously overexpressed proteins in the ER. The use of a reducible cross-linker, DSP, markedly improved the ability to detect noncovalent interactions of PDI, BiP and GRP94 with newly-synthesized thyroglobulin. Under normal conditions, GRP94 was found to associate transiently with early Tg folding intermediates, displaying interaction kinetics similar to those reported for another ER chaperones of BiP.

• Journal of Life Science
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• v.20 no.12
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• pp.1820-1828
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• 2010

Cobalt(II) chloride, a chemical compound with the formula$CoCl_2$, has been widely used in the treatment of anemia, as a chemical agent for the induction of hypoxia in cell cultures, and is known to activate hypoxic signaling. However, excessive exposure to cobalt is associated with several clinical conditions, including asthma, pneumonia, and hematological abnormalities, and can lead to tissue and cellular toxicity. It is also known to induce apoptosis. One of the questions was that of whether $CoCl_2$ might induce apoptosis via endoplasmic reticulum (ER) stress in neurons. To address this question, first, the level of DNA fragmentation was measured for assay of apoptotic rates using $CoCl_2$ with neuron PC12 cells. After confirmation of apoptosis inductions, under the same conditions, the expression levels of ER stress associated factors [ER chaperones Bip, calnexin, ERp72, ERp29, PDI, and ER membrane kinases (IRE1, ATF6, PERK)] were examined by RT-PCR and Western blotting. These results indicated that apoptosis is induced through activation of ER membrane kinases via ER stress. In conclusion, during induction of apoptosis through $CoCl_2$-induced hypoxia in neuron PC12 cells, ER membrane kinase of IRE1 was dominantly up-expressed, and, consecutively, TRAF2, which has been suggested to be one of the links connecting apoptosis and ER stress, was strongly up-expressed.

• Molecules and Cells
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• v.47 no.1
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• pp.100004.1-100004.11
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• 2024

Insulin is essential for maintaining normoglycemia and is predominantly secreted in response to glucose stimulation by β-cells. Incretin hormones, such as glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide, also stimulate insulin secretion. However, as obesity and type 2 diabetes worsen, glucose-dependent insulinotropic polypeptide loses its insulinotropic efficacy, whereas GLP-1 receptor (GLP-1R) agonists continue to be effective owing to its signaling switch from Gs to Gq. Herein, we demonstrated that endoplasmic reticulum (ER) stress induced a transition from Gs to Gq in GLP-1R signaling in mouse islets. Intriguingly, chemical chaperones known to alleviate ER stress, such as 4-PBA and TUDCA, enforced GLP-1R's Gq utilization rather than reversing GLP-1R's signaling switch induced by ER stress or obese and diabetic conditions. In addition, the activation of X-box binding protein 1 (XBP1) or activating transcription factor 6 (ATF6), 2 key ER stress-associated signaling (unfolded protein response) factors, promoted Gs utilization in GLP-1R signaling, whereas Gq employment by ER stress was unaffected by XBP1 or ATF6 activation. Our study revealed that ER stress and its associated signaling events alter GLP-1R's signaling, which can be used in type 2 diabetes treatment.

• Animal cells and systems
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• v.10 no.3
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• pp.115-120
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• 2006

Parkinson's disease (PD) is a neurodegenerative disease caused by selective degeneration of dopaminergic neurons in the substantia nigra. Mutations in ${\alpha}$-synuclein have been causally linked to the pathogenesis of hereditary PD. In addition, it is a major component of Lewy body found in the brains of sporadic cases as well. In the present study, we examined whether overexpression of wild type or PD-related mutant ${\alpha}$-synuclein induces unfolded protein response (UPR) and triggers the known signaling pathway of the resulting endoplasmic reticulum (ER) stress in SH-SY5Y cells. Overexpression of wild type, A30P, and A53T ${\alpha}$-synuclein all induced XBP-1 mRNA splicing, one of the late stage UPR events. However, activation of ER membrane kinases and upregulation of ER or cytoplsmic chaperones were not detected when ${\alpha}$-synuclein was overexpressed. However, basal level of cytoplsmic calcium was elevated in ${\alpha}$-synuclein-expressing cells. Our observation suggests that overexpression of ${\alpha}$-synuclein induces UPR independent of the known ER membrane kinase-mediated signaling pathway and induces ER stress by disturbing calcium homeostasis.

• Animal cells and systems
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• v.7 no.3
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• pp.271-277
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• 2003

Regulation of endoplasmic reticulum(ER) chaperone, ERp29, in traumatized rat spinal cord was investigated. Compared to the control, ERp29 expression was down-regulated at the lesion site 1 d after spinal cord injury. However, ERp29 expression gradually increased from 3 d after the injury and peaked remarkably after 7 d. Two ER chaperones (GRP94 and BiP) showed constantly strong expression levels 1 d after spinal cord injury while the expression levels of the other two (calnexin and PDI) were unchanged. In the case of ERp72, its expression level was increased 1 d after the injury and gradually decreased thereafter. This study suggests that ERp29 expression in the spinal cord after traumatic injury might be associated with the posttraumatic neural survival, playing a role as a molecular chaperone.

• Toxicological Research
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• v.24 no.2
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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.

• International Journal of Industrial Entomology and Biomaterials
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• v.7 no.2
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• pp.133-137
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• 2003

Cells respond to an accumulation of unfolded proteins in the endoplasmic reticulum (ER) by increasing transcription of genes encoding molecular chaperones and folding enzymes. The information is transmitted from the ER lumen to the nucleus by intracellular signaling pathway, called the unfolded protein response (UPR). In Saccharomyces cerevisiae, such induction is mediated by the cis-acting unfolded response element (UPRE) which has been thought to be recognized by Hac1p transcription factor. We cloned the ATFC gene showing similarity with Hac1p, and then examined to determine whether ATFC gene product specifically binds to UPRE by electrophoretic mobility shift assays. ATFC gene product displayed appreciable binding ${to ^{32}}P-labelled$ UPRE. Therefore, we concluded that ATFC represents a major component of the putative transcription factor responsible for the UPR leading to the induction of ER-localized stress proteins.

• YAKHAK HOEJI
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• v.50 no.4
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• pp.263-267
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• 2006

CD1d is an unique antigen presenting molecule which provides antigenic repertoires to NKT cells. To examine molecules required for CD1d antigen presentation, we determined an interaction between CD1d and several endoplasmic reticulum (ER) resident molecular chaperones by co-immunoprecipitation. Results indicated that calnexin and calreticulin seem to be bound to mouse CD1d, but TAP and tapasin do not bind. Further, we screened an yeat two hybrid system to identify proteins that help mouse CD1d transportation in the cytosol. We found that two proteins, heat shock protein a sub-unit $(Hsp90{\alpha})$ and protein kinase C and casein kinase substrate in neurons 3 (PACSIN-3), interact with CD1d. Future study will be focus on the role of these molecules during the CD1d antigen presentation.

• Journal of Physiology & Pathology in Korean Medicine
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• v.23 no.6
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• pp.1368-1378
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• 2009

Unfolded protein response (UPR) is an important genomic response to endoplasmic reticulum (ER) stress. The ER response is characterized by changes in specific proteins, induction of ER chaperones and degradation of misfolded proteins. Also, the pathogenesis of several diseases like Alzheimer's disease, neuronal degenerative diseases, and diabetes reveal the role of ER stress as one of the causative mechanisms. Borneolum has been used for neuronal disease in oriental medicine. In the present study, the protective effect of borneolum on thapsigargin-induced apoptosis in rat C6 glial cells. Treatment with C6 glial cells with 5 uM thapsigargin caused the loss of cell viability, and morphological change, which was associated with the elevation of intracellular $Ca^{++}$ level, the increase in Grp78 and CHOP and cleavage of pro-caspase 12 Furthermore, thapsigargin induced Grp98, XBP1, and ATF4 protein expression in C6 glial cells. Borneolum reduced thapsigargin-induced apoptosis through ER pathways. In the ER pathway, borneolum attenuated thapsigargin-induced elevations in Grp78, CHOP, ATF4, and XBP1 as well as reductions in pro-caspase 12 levels. Also, our data showed that borneolum protected thapsigargin-induced cytotoxicity in astrocytes from rat (P3) brain. Taken together, our data suggest that borneolum is neuroprotective against thapsigargin-induced ER stress in C6 glial cells and astrocytes. Accordingly, borneolum may be therapeutically useful for the treatment of thapsigargin-induced apoptosis in central nervous system.

• Biomolecules & Therapeutics
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• v.17 no.2
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• pp.162-167
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• 2009

Adipocytes undergo adipocyte stress in the excessive presence of lipid. Adipocyte stress accompanies the typical signs of endoplasmic reticulum (ER) stress: unfolded protein response and overexpression of molecular chaperones. Apoptotic induction in adipocytes is known as a good strategy for treating obesity. The drug "tunicamycin" was tested for its therapeutic potential in inducing apoptosis on differentiating adipocytes of 3T3-L1. When the 3T3-L1 cells, stimulated for adipogenesis, were treated with tunicamycin, they showed typical ER stress symptoms. Despite progression in ER stress, however, the differentiated 3T3-L1 hardly proceeded to apoptosis based on the CHOP protein expression and FACS analysis. This is very different from C2C12, the myogenic counterpart of 3T3-L1, which showed significant apoptosis along with ER stress. This study also characterizes a potential mechanism whereby adipocyte may avoid apoptosis to sustain the pathological state of obesity. The level of GRP94 expression significantly upholds in 3T3-L1 under tunicamycin treatment compared to preadipocytes and C2C-12. When GRP94 expression was inhibited by siRNA, 3T3-L1 showed a higher level of CHOP expression compared to C2C12 cells. In conclusion, adipocytes exert an anti-apoptotic mechanism under ER stress caused by tunicamycin; thus, apoptotic induction in adipocyte is not a viable anti-obesity option. The unusual level of GRP94 may serve as a key role whereby adipocytes reach to the obesity level circumventing the apoptosis.