• Molecules and Cells
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• v.42 no.12
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• pp.840-849
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• 2019

The spatiotemporal mitotic processes are controlled qualitatively by phosphorylation and qualitatively by ubiquitination. Although the SKP1-CUL1-F-box protein (SCF) complex and the anaphase-promoting complex/cyclosome (APC/C) mainly mediate ubiquitin-dependent proteolysis of mitotic regulators, the E3 ligase for a large portion of mitotic proteins has yet to be identified. Here, we report c-Cbl as an E3 ligase that degrades DDA3, a protein involved in spindle dynamics. Depletion of c-Cbl led to increased DDA3 protein levels, resulting in increased recruitment of Kif2a to the mitotic spindle, a concomitant reduction in spindle formation, and chromosome alignment defects. Furthermore, c-Cbl depletion induced centrosome over-duplication and centriole amplification. Therefore, we concluded that c-Cbl controls spindle dynamics and centriole duplication through its E3 ligase activity against DDA3.

• Molecules and Cells
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• v.26 no.3
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• pp.285-290
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• 2008

Estrogen-induced proliferation in estrogen receptor (ER)-positive breast cancer cells is primarily mediated through two distinct intracellular receptors, $ER{\alpha}$ and $ER{\beta}$. Although tumor necrosis factor alpha ($TNF{\alpha}$) and $E2/ER{\alpha}$ are known to exert opposing effects on cell proliferation in MCF-7 cells, the mechanism by which $TNF{\alpha}$ antagonizes $E2/ER{\alpha}$-mediated cell proliferation is not well understood. The present study suggests that reduced cell survival in response to $TNF{\alpha}$ treatment in MCF-7 cells may be associated with the down-regulation of $ER{\alpha}$ protein. The decrease in $ER{\alpha}$ protein level was accompanied by an inhibition of $ER{\alpha}$ gene transcription. Cell viability was decreased synergistically by the combined treatment with $ER{\alpha}$-siRNA and $TNF{\alpha}$. Furthermore, pretreatment of cells with the PI3-kinase (PI3K)/ Akt inhibitor, LY294002, markedly enhanced $TNF{\alpha}$-induced down-regulation of the $ER{\alpha}$ protein, suggesting that the PI3K/Akt pathway might be involved in control of the $ER{\alpha}$ level. Moreover, down-regulation of $ER{\alpha}$ by $TNF{\alpha}$ was not inhibited in cells that were pretreated with the proteasome inhibitors, MG132 and MG152, which suggests that proteasome-dependent proteolysis does not significantly influence $TNF{\alpha}$-induced down-regulation of $ER{\alpha}$ protein. In contrast, the effect of the PI3K/Akt inhibitor on $ER{\alpha}$ was blocked in cells that were treated with LY294002 in the presence of the proteasome inhibitors. Collectively, our findings show that the $TNF{\alpha}$ may partly regulate the growth of MCF-7 breast cancer cells through the down-regulation of $ER{\alpha}$ expression, which is primarily mediated by a PI3K/Akt signaling.

• Animal cells and systems
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• v.5 no.4
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• pp.267-274
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• 2001

Calpains are a family of cysteine proteases existing primarily in two forms designated by the $Ca^{2+}$ concentration needed for activation in vitro, $\mu$-calpain (calpain-I) and m-calpain (calpain-II). The physiologica1 roles of calpains remain unclear. Many groups have proposed a role for calpains In apoptosis, but their patterns of activation are not well characterized. Calpains have been implicated in neutrophil apoptosis, glucocorticoid-induced thymocyte apoptosis, as well as many other apoptotic pathways. Calpain activation in apoptosis is usually linked upstream or downstream to caspase activation, or in a parallel pathway alongside caspase activation. Calpains have been suggested to be involved in DNA fragmentation (via endonuclease activation), but also as effector proteases that cleave cellular proteins involved in DNA repair, membrane associated proteins and other homeostatic regulatory proteins. Recently, our laboratory demonstrated $\mu$-calpain activation in NAD(P)H: quinone oxidoreducatse 1 (NQO1)-expressing cells after exposure to $\beta$-lapachone, a novel quinone and potential chemo- and radio-therapeutic agent. Increased cytosolic $Ca^{2+}$ in NQO1-expressing cells after $\beta$-lapachone exposures were shown to lead to $\mu$-calpain activation. In turn, $\mu$-calpain activation was important for substrate proteolysis and DNA fragmentation associated with apoptosis. Upon activation, $\mu$-calpain translocated to the nucleus where it could proteolytically cleave PARP and p53. We provided evidence that $\beta$-lapachone-induced, $\mu$-calpain stimulated, apoptosis did not involve any of the known caspases; known apoptotic caspases were not activated after $\beta$-lapachone treatment of NQO1-expressing cells, nor did caspase inhibitors have any effect on $\beta$-1apachone-induced cell death. Elucidation of processes by which $\beta$-1apachone-stimulated $\mu$-calpain activation and calpains ability to activate endonucleases and induce apoptosis independent of caspase activity will be needed to further develop/modulate $\beta$-lapachone for treatment of human cancers that over-express NQO1.

• Proceedings of the Microbiological Society of Korea Conference
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• 2004.05a
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• pp.65-67
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• 2004

O-GlcNAcylation of p53 has been already identified and reported, but the function of O-GlcNAc on p53 has not been studied well. In this report, the general function of O-GlcNAc modification on p53 has been investigated using mouse fibroblast cell, L929. When streptozotocin (STZ), a non-competitive O-GlcNAcase inhibitor was treated to L929, O-GlcNAc modification level was dramatically increased on nucleocytoplasmic proteins, including p53. Because it has been already reported that $TNF\alpha$ induced the production of p53 in L929, $TNF\alpha$ was treated to obtain more p53. Approximately two times more amount of p53 was found from the cells treated STZ and $TNF\alpha$ simultaneously compared to the cell treated $TNF\alpha$ alone. The p53 increment in the presence of STZ was not caused by the induction of p53 gene expression. When new production of p53 induced by the $TNF\alpha$ was inhibited by the treatment of cycloheximide, O-GlcNAc modification decreased and phosphorylation increased on pre-existing p53 after $TNF\alpha$ treatment. But in the presence of STZ and $TNF\alpha$ at the same time, more O-GlcNAcylation occurred on p53, The level of ubiquitination on p53 was also reduced in the presence of STZ. Approximately three times less amount of Mdm2 bound to this hyperglycosylated p53. From this result it might be concluded that treatment of STZ to inhibit O-GlcNAcase increased O-GlcNAc modification level on p53 and the increment of O-GlcNAc modification stabilized p53 from ubiquitin proteolysis system.

• Journal of Korean Medicine Rehabilitation
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• v.33 no.3
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• pp.17-32
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• 2023

Objectives The purpose of this study was to investigate the antioxidant, anti-inflammatory and cartilage regeneration effects of Euiiin-tang water extract (EIIT) in the treatment of monosodium iodoacetate (MIA)-induced osteoarthritis in rats. Methods Animal models were divided into five groups. The normal group didn't do any treatments causing osteoarthritis. The control group was orally administerd distilled water instead of the drug, the positive control group used indomethacin 5 mg/kg, the EIIT 100 group used EIIT 100 mg/kg and the EIIT 200 group used EIIT 200 mg/kg, and seven rats were placed per group. We administered drug to rats for 2 weeks and analyzed oxidative stress-related proteins in joint tissue. Inflammation mediators and inflammatory cytokines induced by the activity of inflammation-related proteins were analyzed. In addition, the expression of anti-inflammatory cytokines and collagen-related factors were analyzed, and H&E staining and Safranin-O staining were performed to see the effect on histopathological changes. Results 1) Oxidative stress-related proteins were significantly reduced. 2) Inflammationrelated proteins, inflammatory mediators and inflammatory cytokines were significantly reduced. 3) Anti-inflammatory cytokines were significantly increased. 4) Collagen proteolysis factors significantly decreased, and collagen degradation inhibitory factor was significantly increased. 5) EIIT administration significantly reduced cartilage degeneration and deformation in H&E staining, and reduced proteoglycan destruction in Safranin-O staining. Conclusions From the above experimental results, it judges that Euiiin-tang has antioxidant, anti-inflammatory, and cartilage regeneration effects on osteoarthritis in rats induced by MIA.

• Molecules and Cells
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• v.21 no.2
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• pp.218-223
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• 2006

The effect of poly(ADP-ribosyl)ation on the stability of p53 in SK-HEP1 cells treated with UV light was examined. Intracellular levels of p53 increased in cells treated with a low dose of UV light ($20J/m^2$), whereas they increased but then declined after a higher dose of UV ($100J/m^2$). Intracellular levels of p53 in the UV treated SK-HEP1 cells were dependent on the UV dose. Use of proteasome inhibitors revealed that p53 is degraded by proteasomal proteolysis after high doses of UV light. We present evidence that, at low doses, poly(ADP-ribose)polymerase (PARP) poly(ADP-ribosyl) ates p53 and protects it from proteasomal degradation before caspase-3 is activated, whereas at high doses the cells undergo UV induced apoptosis and PARP is cleaved by caspase-3 before it can protect p53 from degradation. Destabilization of p53 by cleavage of PARP may be important in cell fate decision favoring apoptosis.

• BMB Reports
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• v.54 no.6
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• pp.305-310
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• 2021

Cereblon (CRBN) is a multi-functional protein that acts as a substrate receptor of the E3 ligase complex and a molecular chaperone. While CRBN is proposed to function in mitochondria, its specific roles are yet to be established. Here, we showed that knockdown of CRBN triggers oxidative stress and calcium overload in mitochondria, leading to disruption of mitochondrial membrane potential. Notably, long-term CRBN depletion using PROteolysis TArgeting Chimera (PROTAC) induced irreversible mitochondrial dysfunction, resulting in cell death. Our collective findings indicate that CRBN is required for mitochondrial homeostasis in cells.

• Journal of Animal Science and Technology
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• v.65 no.6
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• pp.1151-1168
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• 2023

Tenderness and taste characteristics of meat are the key determinants of the meat choices of consumers. This review summarizes the contemporary research on the molecular mechanisms by which postmortem aging of meat improves the tenderness and taste characteristics. The fundamental mechanism by which postmortem aging improves the tenderness of meat involves the operation of the calpain system due to apoptosis, resulting in proteolytic enzyme-induced degradation of cytoskeletal myofibrillar proteins. The improvement of taste characteristics by postmortem aging is mainly explained by the increase in the content of taste-related peptides, free amino acids, and nucleotides produced by increased hydrolysis activity. This review improves our understanding of the published research on tenderness and taste characteristics of meat and provides insights to improve these attributes of meat through postmortem aging.

• Herbal Formula Science
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• v.29 no.4
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• pp.239-252
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• 2021

Objectives : This study is to effect of improving muscle atrophy through water extract on the solid-phase fermentation extraction with Phellinus linteus of discarded Schisandra chinensis in an atorvastatin-induced atrophy C2C12 cell. Methods : C2C12 myoblast were differentiated into myotube by 2% horse serum medium for 6 days, and then treated solid-phase fermentation(S-P) extract at different concentrations for 24h. To investigate the effect of S-P extract on the induction of muscle atrophy and expression of atrophy-related genes and apoptosis in differentiated C2C12 myotubes using a GSH, ROS, real-time PCR, western blots analysis. Results : As a result of treatment with atorvastatin at concentrations of 5, 10, and 20 uM on the 6th day of differentiation in C2C12 myotube cells, it was confirmed that the cell morphology was damaged in a concentration-dependent manner, and the length and thickness of the myotube also decreased in a concentration-dependent manner. Treatment with S-P extract (50, 100 and 200 ㎍/㎖) increased of GSH and inhibited ROS in the atorvastatin-induced muscle atrophy cell model at a concentration that did not induce toxicity. In addition, it was confirmed that it has an effect on muscle reduction by inhibiting apoptosis of muscle cells as well as being involved in protein production and degradation of muscle cells. Conclusions : Atorvastatin-induced atrophy C2C12 cell, S-P extract activates related to differentiation/generation and proteolysis, and inhibits cell death of atrophy in C2C12 cell. Based on this, it is necessary to prove its effectiveness through animal models and human application test, but it is considered to be discarded Schisandra chinensis can present the potential for development as a recycling industrial material.

• Journal of Physiology & Pathology in Korean Medicine
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• v.16 no.1
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• pp.172-180
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• 2002

Polychlorinated biphenyls(PCBs) are large scale industrial chemicals which are using in diverse applications. The goal of this study was to determine if exposure to 2,2',5,5'-tetrachlorobiphenyl (PCB 52) leads to an increase in the production of active oxidants, and subsequently promotes apoptosis of neuronal SK-N-MC cells. Reactive oxygen species (ROS) formation was examined in SK-N-MC cells after treatment of PCB 52 by concentrations and incubation times, respectively. It showed that the rate of ROS production in the cells was increased in a does-dependent manner to 45 min, followed by a return towards control levels after 120 min treatment. We also examined the association of PCB-induced apoptosis with the modulation of biomakers of oxidative damage to lipids (malondialdehyde [MDA]) in SK-N-MC cells. Increased MDA was observed in a dose-dependent manner in groups treated with 10, 15, and 20 figJ me of PCB 52 for 24 h. After treatment of PCB 52, the cells did not show any significant change in the rate of Cu/Zn-superoxide dismutase (Cu/Zn-SOD) activity. Whereas, the cells had a two-fold greater rate of change in catalase activity at 20 ㎍/㎖ of PCB 52 for 24 h when compared to control group. Korean Ginseng is one of the most important crude drugs which has been used as a traditional Oriental medicine. We next investigated protective effect of extracts of ginseng on cytotoxicity induced by PCB 52 in SK-N-MC cells. Pretreatment of SK-N-MC cells with 25-200 μg/ml of ginseng were reduced cell death in a dose-dependent manner in PCB 52-treated cells. To examine the sensitivity of beta-catenin to ginseng, the protective effect of a range of ginseng concentrations was examined in SK-N-MC cells treated with PCB 52. The result demonstrated that ginseng efficiently blocked PCB 52 inducible beta-catenin proteolysis in a concentration dependent manner. The ROS formation was also measured in the presences of extract of ginseng and superoxide dismutase (inhibitor of oxygen free radical production). The both SOD (400 U/ml) and ginseng (200 μg/ml) significantly inhibited RDS generation in PCB 52-treated group.