• BMB Reports
• /
• 제38권6호
• /
• pp.709-716
• /
• 2005

Apoptosis and necrosis are distinguished by modality primarily. Here we show an apoptosis occurred instantly, induced by $300\;{\mu}M$ W-7 ((N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride), inhibitor of calmodulin), which demonstrated necrotic modality. As early as 30 min after W-7 addition, apoptotic (sub-diploid) peak could be detected by fluorescence-activated cell sorter (FACS), “DNA ladders” began to emerge also at this time point, activity of caspase-3 elevated obviously within this period. Absence of mitochondrial membrane potential (MMP) reduction and cytochrome c, AIF (apoptosis inducing factor) release, verified that this rapid apoptosis did not proceed through mitochondria pathway. Activation of caspase-12 and changes of other endoplasmic reticulum (ER) located proteins ascertained that ER pathway mediated this necrosis-like apoptosis. Our findings suggest that it is not credible to judge apoptosis by modality. Elucidation of ER pathway is helpful to comprehend the pathology of diseases associated with ER stress, and may offer a new approach to the therapy of cancer and neurodegenerative diseases.

• 생명과학회지
• /
• 제20권12호
• /
• pp.1820-1828
• /
• 2010

PC12 세포에서 $CoCl_2$에 의한 hypoxia 유도는 HIF1 alpha의 상승 발현으로 확인하였다. 이때 apoptosis의 유도는 genomic DNA의 fragmentation과 apoptotic body는 Hoechst 염색으로 확인되었고, ER luminal chaperone의 발현 및 ER stress signal에 관여하는 ER membrane kinase인 IRE1, PERK, ATF6의 발현도 확인되었다. 이들이 apoptosis로 연결되는 고리 역할을 하는 IRE1-XBP1 mRNA splicing, PERK-eIF2 alpha, ATF6 protein cleavage도 반응하는 것으로 확인되었다. 위의 결과는 신경세포의 hypoxia상태는 ER stress signal pathway를 거쳐서 apoptosis가 된다는 것을 증명한 것으로 신경세포의 hypoxia치료를 위한 기초 자료가 될 것으로 생각한다.

• Animal Bioscience
• /
• 제34권6호
• /
• pp.1006-1013
• /
• 2021

Objective: Cells have increased susceptibility to activation of apoptosis when suffering heat stress (HS). An effective supplementation strategy to mimic heat-induced apoptosis of bovine mammary epithelial cells (MECs) is necessary to maintain optimal milk production. This study aimed to investigate possible protective effects of the anti-apoptotic activity of 5-aminolevulinic acid (5-ALA) against HS-induced damage of bovine MECs. Methods: Bovine MECs were pretreated with or without 5-ALA at concentrations of 10, 100, and 500 µM for 24 h followed by HS (42.5℃ for 24 h and 48 h). Cell viability was measured with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. Real-time quantitative polymerase chain reaction and Western blotting were used to explore the regulation of genes associated with apoptosis, oxidative stress, and endoplasmic reticulum (ER) stress genes. Results: We found that 5-ALA induces cytoprotection via inhibition of apoptosis markers after HS-induced damage. Pretreatment of bovine MECs with 5-ALA resulted in dramatic upregulation of mRNA for nuclear factor erythroid-derived 2-like factor 2, heme oxygenase-1, and NAD(P)H quinone oxidoreductase 1, all of which are antioxidant stress genes. Moreover, 5-ALA pretreatment significantly suppressed HS-induced ER stress-associated markers, glucose-regulated protein 78, and C/EBP homologous protein expression levels. Conclusion: 5-ALA can ameliorate the ER stress in heat stressed bovine MEC via enhancing the expression of antioxidant gene.

• 생명과학회지
• /
• 제21권4호
• /
• pp.613-615
• /
• 2011

ERp29는 endoplasmic reticulum (ER) lumen에 존재하는 단백질로 protein disulfide isomerase (PDI) family에 속한다. 비록 관련 연구 결과는 조금 있지만 정확한 생물학적인 기능은 아직 분명하지 않지만, 분비단백질과정과 단백질 folding에 관여하는 것으로 알려 지고 있다. ERp29의 기능 연구를 위하여 yeast two-hybrid screening/GST pull-down assay방법을 사용하여 ERp29-결합단백질인 ADP-ribosylation factor 5 (ARF5)를 동정하였다. 이들의 결합은 정상적인 세포생리상태에서는 결합하지만 ER stress 상태에서는 떨어졌다. 이 결과는 ERp29의 기능 연구를 위하여 하나의 실마리를 제공할 것이다.

• 생명과학회지
• /
• 제28권10호
• /
• pp.1186-1192
• /
• 2018

비만에 의한 근형질세망 스트레스(Endoplasmic stress, ER stress) 발생은 비접힙 단백질의 축적으로 근육내 대사와 근기능 저하를 초래한다. 유산소 운동과 폴리페놀은 비만에 유도된 염증성 사이토카인과 ER stress를 감소시키는 것으로 잘 알려졌다. 그러나 신체활동과 약물에 의한 염증성 사이토카인과 ER stress 변인들의 변화에 선택적인 반응이 있는지에 대한 연구는 이루어지지 않았다. 따라서, 이 연구의 목적은 유산소 운동과 폴리페놀섭취에 따른 골격근내 ER stress 관련 변인에 우선적인 효과가 있는지 비교분석하고자 하였다. 이 연구의 목적을 위해 (1) 정상식이 그룹, (2) 60% 고지방식이 그룹, (3) 고지방식와 레스베라트롤 25 mg/kg 그룹, (4) 고지방식이와 크리신 50 mg/kg 투여 그룹, (5) 고지방식이와 운동 그룹으로 나누어 실시하였다. 레스베라트롤과 크리신 그룹은 16주간 경구 투여 하였고, 운동 그룹은 1일 40-60분간 10-14 m/min의 속도로 주 4일, 총 16주간 트레드밀 운동을 수행하였다. 운동그룹에서 $IRE1{\alpha}$, BIP/GRP78는 고지방 식이 그룹과 비교하여 유의하게 낮아졌다(p<0.05). 고지방식이와 레스베라트롤 투여 그룹에서 ATF6, $IRE1{\alpha}$, BIP/GRP78는 고지방식이 그룹과 비교하여 유의하게 낮아졌다(p<0.05). 고지방식이와 크리신 투여 그룹은 고지방식이 그룹과 비교하여 ATF6가 유의하게 낮아졌다(p<0.05). 이러한 결과는 유산소 운동과 레스베라트롤, 크리신 섭취에 의해 ER stress 관련 변인을 조절할 수 있다. 그러나 유산소 운동은 고지방 식이에 의해 유도되는 ER stress 개선이 더욱 효과적인 것으로 나타났다.

• BMB Reports
• /
• 제53권2호
• /
• pp.88-93
• /
• 2020

Cisplatin is a widely used anti-cancer agent. However, the effectiveness of cisplatin has been limited by the commonly developed drug resistance. This study aimed to investigate the potential effects of endoplasmic reticulum (ER) stress to overcome drug resistance using the cisplatin-resistant A2780/CisR ovarian cancer cell model. The synthetic chalcone derivative (E)-3-(3,5-dimethoxyphenyl)-1-(2-methoxyphenyl)prop-2-en-1-one (named DPP23) is an ER stress inducer. We found that DPP23 triggered apoptosis in both parental cisplatin-sensitive A2780 and cisplatin-resistant A2780/CisR ovarian cancer cells due to activation of reactive oxygen species (ROS)-mediated unfolded protein response (UPR) pathway in the endoplasmic reticulum. This result suggests that ROS-mediated UPR activation is potential in overcoming drug resistance. DPP23 can be used as a target pharmacophore for the development of novel chemotherapeutic agents capable of overcoming drug resistance in cancer cells, particularly ovarian cancer cells.

• Animal Bioscience
• /
• 제35권1호
• /
• pp.22-28
• /
• 2022

Objective: Endoplasmic reticulum (ER) stress engages the unfolded protein response (UPR) that serves as an important mechanism for modulating hepatic fatty acid oxidation and lipogenesis. Chronic fasting in mice induced the UPR activation to regulate lipid metabolism. However, there is no direct evidence of whether negative energy balance (NEB) induces ER stress in the liver of cows. This study aimed to elucidate the relationship between the NEB attributed to feed deprivation and ER stress in bovine hepatocytes. Methods: Blood samples and liver biopsy tissues were collected from 6 non-lactating cows before and after their starvation for 48 h. The blood non-esterified fatty acids (NEFA), β-hydroxybutyric acid (BHBA) and glucose level were analyzed. Real-time quantitative polymerase chain reaction and Western blotting were used to explore the regulation of genes associated with UPR and lipid metabolism. Results: The starvation increased the plasma BHBA and NEFA levels and decreased the glucose level. Additionally, the starvation caused significant increases in the mRNA expression level of spliced X-box binding protein 1 (XBP1s) and the protein level of phosphorylated inositol-requiring kinase 1 alpha (p-IRE1α; an upstream protein of XBP1) in the liver. The mRNA expression levels of peroxisome proliferator-activated receptor alpha and its target fatty acid oxidation- and ketogenesis-related genes were significantly upregulated by the starvation-mediated NEB. Furthermore, we found that the mRNA expression levels of lipogenic genes were not significantly changed after starvation. Conclusion: These findings suggest that in the initial stage of NEB in dairy cows, the liver coordinates an adaptive response by activating the IRE1 arm of the UPR to enhance ketogenesis, thereby avoiding a fatty liver status.

• BMB Reports
• /
• 제48권8호
• /
• pp.445-453
• /
• 2015

Endoplasmic Reticulum (ER) is an organelle where most secretory and membrane proteins are synthesized, folded, and undergo further maturation. As numerous conditions can perturb such ER function, eukaryotic cells are equipped with responsive signaling pathways, widely referred to as the Unfolded Protein Response (UPR). Chronic conditions of ER stress that cannot be fully resolved by UPR, or conditions that impair UPR signaling itself, are associated with many metabolic and degenerative diseases. In recent years, Drosophila has been actively employed to study such connections between UPR and disease. Notably, the UPR pathways are largely conserved between Drosophila and humans, and the mediating genes are essential for development in both organisms, indicating their requirement to resolve inherent stress. By now, many Drosophila mutations are known to impose stress in the ER, and a number of these appear similar to those that underlie human diseases. In addition, studies have employed the strategy of overexpressing human mutations in Drosophila tissues to perform genetic modifier screens. The fact that the basic UPR pathways are conserved, together with the availability of many human disease models in this organism, makes Drosophila a powerful tool for studying human disease mechanisms. [BMB Reports 2015; 48(8): 445-453]

• Molecules and Cells
• /
• 제47권1호
• /
• pp.100001.1-100001.8
• /
• 2024

In eukaryotes, a primary protein quality control (PQC) process involves the destruction of conformationally misfolded proteins through the ubiquitin-proteasome system. Because approximately one-third of eukaryotic proteomes fold and assemble within the endoplasmic reticulum (ER) before being sent to their destinations, the ER plays a crucial role in PQC. The specific functions and biochemical roles of several E3 ubiquitin ligases involved in ER-associated degradation in mammals, on the other hand, are mainly unknown. We identified 2 E3 ligases, ubiquitin protein ligase E3 component N-recognin 1 (UBR1) and ubiquitin protein ligase E3 component N-recognin 2 (UBR2), which are the key N-recognins in the N-degron pathway and participate in the ER stress response in mammalian cells by modulating their stability. Cells lacking UBR1 and UBR2 are hypersensitive to ER stress-induced apoptosis. Under normal circumstances, these proteins are polyubiquitinated through Lys48-specific linkages and are then degraded by the 26S proteasome. In contrast, when cells are subjected to ER stress, UBR1 and UBR2 exhibit greater stability, potentially as a cellular adaptive response to stressful conditions. Although the precise mechanisms underlying these findings require further investigation, our findings show that cytoplasmic UBR1 and UBR2 have anti-ER stress activities and contribute to global PQC in mammals. These data also reveal an additional level of complexity within the mammalian ER-associated degradation system, implicating potential involvement of the N-degron pathway.

• 생명과학회지
• /
• 제17권7호통권87호
• /
• pp.1023-1025
• /
• 2007

Ajuba is a number of proteins containing cytosolic LIM domain. Its function may provide a new pathway whereby cell-cell adhesive events are transmitted to the nucleus to regulate cell proliferation and differentiation decisions. Here, Ajuba gene expression was investigated its molecular properties associated with endoplasmic reticulum (ER) stresses (tunicamycin, DTT, A23187 and BFA) which induced remarkable ex-pression of Ajuba mRNA. The mRNA half life of Ajuba was also determined, its half life of Ajuba mRNA in FRTL-5 cells was approximately 2 hr after the initial translation. Although the obvious bioligical function of Ajuba is not clear, on the base of the results, Ajuba gene expression is deeply associated with ER stresses.