• Biomolecules & Therapeutics
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• 제25권4호
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• pp.396-403
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• 2017

Under normal, non-stressed conditions, intracellular p53 is continually ubiquitinated by MDM2 and targeted for degradation. However, in response to severe genotoxic stress, p53 protein levels are markedly increased and apoptotic cell death is triggered. Inhibiting the ubiquitination of p53 under conditions where DNA damage has occurred is therefore crucial for preventing the development of cancer, because if cells with severely damaged genomes are not removed from the population, uncontrolled growth can result. However, questions remain about the cellular mechanisms underlying the regulation of p53 stability. In this study, we show that p53-inducible gene 3 (PIG3), which is a transcriptional target of p53, regulates p53 stability. Overexpression of PIG3 stabilized both endogenous and transfected wild-type p53, whereas a knockdown of PIG3 lead to a reduction in both endogenous and UV-induced p53 levels in p53-proficient human cancer cells. Using both in vivo and in vitro ubiquitination assays, we found that PIG3 suppressed both ubiquitination- and MDM2-dependent proteasomal degradation of p53. Notably, we demonstrate that PIG3 interacts directly with MDM2 and promoted MDM2 ubiquitination. Moreover, elimination of endogenous PIG3 in p53-proficient HCT116 cells decreased p53 phosphorylation in response to UV irradiation. These results suggest an important role for PIG3 in regulating intracellular p53 levels through the inhibition of p53 ubiquitination.

• KSBB Journal
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• 제24권3호
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• pp.217-226
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• 2009

p53은 전사인자로서 세포의 사멸이나 세포주기 조절 등 다양한 세포 활성을 보이기 때문에 일반적인 환경에서는 매우 낮은 수준으로 단백질 양이 확인된다. p53의 단백질 양과 활성은 다양한 세포 내 신호에 의하여 이루어지는 후전사 변형을 통하여 조절 받는다. 이중 유비퀴틴화는 세포 내에서 p53 단백질의 발현 수준이 낮게 유지되는 것이 가능하게 하는 대표적인 기전이다. 이러한 기전을 일으키는 대표적인 p53의 E3 ligase로는 mdm2, Pirh2, COP1, ARF-BP1 등이 보고되어 있으며, 각각 negative feedback loop나 다른 기전을 통하여 p53 단백질의 분해를 유도하여 세포의 항상성을 조절한다. 이 밖에도 p53은 mdm2나 WWP1, UBC13, MSL2와 같은 E3 ligase로 인해서 모노 유비퀴틴화 되고, p53의 세포 내 위치가 조절되어 전사인자로서의 활성이 억제된다. p53의 세포 내 위치와는 관계없이 p53의 전사인자로써의 활성 또한 아세틸화와 유비퀴틴화의 경쟁적 반응으로 인해 조절 될 수 있다. E4F1에 의한 유비퀴틴화는 세포주기와 관련된 유전자의 발현을 증가시키되 세포사멸 관련 유전자의 발현은 감소시키는 것으로 보아 p53의 수많은 downstream gene의 발현 또한 유비퀴틴화를 통해 조절 될 수 있음이 제시되었다. 앞으로의 연구는 신규 E3 ligase에 의한 p53의 유비퀴틴화 기전 연구 뿐 아니라 이와 관련된 다른 변형과의 관계에 대한 연구 또한 매우 중요하게 부각되어 질 것으로 예상된다.

• BMB Reports
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• 제49권11호
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• pp.598-606
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• 2016

ARF is an alternative reading frame product of the INK4a/ARF locus, inactivated in numerous human cancers. ARF is a key regulator of cellular senescence, an irreversible cell growth arrest that suppresses tumor cell growth. It functions by sequestering MDM2 (a p53 E3 ligase) in the nucleolus, thus activating p53. Besides MDM2, ARF has numerous other interacting partners that induce either cellular senescence or apoptosis in a p53-independent manner. This further complicates the dynamics of the ARF network. Expression of ARF is frequently disrupted in human cancers, mainly due to epigenetic and transcriptional regulation. Vigorous studies on various transcription factors that either positively or negatively regulate ARF transcription have been carried out. However, recent focus on posttranslational modifications, particularly ubiquitination, indicates wider dynamic controls of ARF than previously known. In this review, we discuss the role and dynamic regulation of ARF in senescence and cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제15권5호
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• pp.2313-2318
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• 2014

Peroxisome proliferator-activated receptor gamma (PPAR-${\gamma}$), a ligand-dependent nuclear transcription factor, has been found to widely exist in tumor tissues and plays an important role in affecting tumor cell growth. In this study, we investigated the effect of PPAR-${\gamma}$ on aspects of the cervical cancer malignant phenotype, such as cell proliferation and apoptosis. Cell growth assay, Western blotting, Annexin V and flow cytometry analysis consistently showed that treatment with troglitazone (TGZ, a PPAR-${\gamma}$ agonist) led to dose-dependent inhibition of cervical cancer cell growth through apoptosis, whereas T0070907 (another PPAR-${\gamma}$ antagonist) had no effect on Hela cell proliferation and apoptosis. Furthermore, we also detected the protein expression of p53, p21 and Mdm2 to explain the underlying mechanism of PPAR-${\gamma}$ on cellular apoptosis. Our work, finally, demonstrated the existence of the TGZ-PPAR-${\gamma}$-p53 signaling pathway to be a critical regulator of cell apoptosis. These results suggested that PPAR-${\gamma}$ may be a potential therapeutic target for cervical cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제16권9호
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• pp.3723-3727
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• 2015

The p53 tumor suppressor protein is a principal mediator of growth arrest, senescence, and apoptosis in response to a broad array of cellular damage. p53 is a substrate for the ubiquitin-proteasome system, however, the ubiquitin-conjugating enzymes (E2s) involved in p53 ubiquitination have not been well studied. UBE2Q1 is a novel E2 ubiquitin conjugating enzyme gene. Here, we investigated the effect of UBE2Q1 overexpression on the level of p53 in the MDA-MB-468 breast cancer cell line as well as the interaction between UBE2Q1 and p53. By using a lipofection method, the p53 mutated breast cancer cell line, MDA-MB-468, was transfected with the vector pCMV6-AN-GFP, containing UBE2Q1 ORF. Western blot analysis was employed to verify the overexpression of UBE2Q1 in MDA-MB-468 cells and to evaluate the expression level of p53 before and after cell transfection. Immunoprecipitation and GST pull-down protocols were used to investigate the binding of UBE2Q1 to p53. We established MDA-MB-468 cells that transiently expressed a GFP fusion proteins containing UBE2Q1 (GFP-UBE2Q1). Western blot analysis revealed that levels of p53 were markedly lower in UBE2Q1 transfected MDA-MB-468 cells as compared with control MDA-MB-468 cells. Both in vivo and in vitro data showed that UBE2Q1 co-precipitated with p53 protein. Our data for the first time showed that overexpression of UBE2Q1can lead to the repression of p53 in MDA-MB-468 cells. This repression of p53 may be due to its UBE2Q1 mediated ubiquitination and subsequent proteasome degradation, a process that may involve direct interaction of UBE2Q1with p53.

• The Korean Journal of Physiology and Pharmacology
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• 제20권5호
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• pp.487-498
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• 2016

Leptin, an adipokine predominantly produced from adipose tissue, is well known to induce tumor growth. However, underlying molecular mechanisms are not established yet. While p53 has long been well recognized as a potent tumor suppressor gene, accumulating evidence has also indicated its potential role in growth and survival of cancer cells depending on experimental environments. In the present study, we examined if p53 signaling is implicated in leptin-induced growth of cancer cells. Herein, we demonstrated that leptin treatment significantly increased p53 protein expression in both hepatic (HepG2) and breast (MCF-7) cancer cells without significant effect on mRNA expression. Enhanced p53 expression by leptin was mediated via modulation of ubiquitination, in particular ubiquitin specific protease 2 (USP2)-dependent manner. Furthermore, gene silencing of p53 by small interfering RNA (siRNA) suppressed leptin-induced growth of hepatic and breast cancer cells, indicating the role of p53 signaling in tumor growth by leptin. In addition, we also showed that knockdown of p53 restored suppression of caspase-3 activity by leptin through modulating Bax expression and prevented leptin-induced cell cycle progression, implying the involvement of p53 signaling in the regulation of both apoptosis and cell cycle progression in cancer cells treated with leptin. Taken together, the results in the present study demonstrated the potential role of p53 signaling in leptin-induced tumor growth.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2004년도 International Meeting of the Microbiological Society of Korea
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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.

• BMB Reports
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• 제51권12호
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• pp.642-647
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• 2018

Ubiquitin-conjugating enzyme E2S (UBE2S), a family of E2 protein in the ubiquitination process, is involved in development of various cancers. However, its role in lung adenocarcinoma, has not been well elucidated. In this report, we attempted to investigate expression and function of UBE2S in lung adenocarcinoma. Up-regulation of UBE2S at mRNA, and protein level, was observed in human cancer tissues and lung adenocarcinoma cells. Higher UBE2S expression correlated with poorer prognosis of lung adenocarcinoma patients. UBE2S expression was efficiently suppressed by lentivirus-mediated shRNA strategy in A549 cells, and UBE2S silencing led to reduced cell proliferation, colony formation, and enhanced apoptosis. Inverse results were observed, in UBE2S over-expressed H1299 cells. Microarray analysis indicated that a large number of genes were regulated by UBE2S, and p53 signaling pathway may be critical, to the role of UBE2S in cancer development. Together, UBE2S could be a potential target for lung adenocarcinoma.

• BMB Reports
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• 제36권1호
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• pp.120-127
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• 2003

The formation of new blood vessels, angiogenesis, is an essential process during development and disease. Angiogenesis is well known as a crucial step in tumor growth and progression. Angiogenesis is induced by hypoxic conditions and regulated by the hypoxia-inducible factor 1 (HIF-1). The expression of HIF-1 correlates with hypoxia-induced angiogenesis as a result of the induction of the major HIF-1 target gene, vascular endothelial cell growth factor (VEGF). In this review, a brief overview of the mechanism of angiogenesis is discussed, focusing on the regulatory processes of the HIF-1 transcription factor. HIF-1 consists of a constitutively expressed HIF-1 beta(HIF-1β) subunit and an oxygen-regulated HIF-1 alpha(HIF-1α) subunit. The stability and activity of HIF-1α are regulated by the interaction with various proteins, such as pVHL, p53, and p300/CBP as well as by post-translational modifications, hydroxylation, acetylation, and phosphorylation. It was recently reported that HIF-1α binds a co-activator of the AP-1 transciption factor, Jab-1, which inhibits the p53-dependent degradation of HIF-1 and enhances the transcriptional activity of HIF-1 and the subsequent VEGF expression under hypoxic conditions. ARD1 acetylates HIF-1α and stimulates pVHL-mediated ubiquitination of HIF-1α. With a growing knowledge of the molecular mechanisms in this field, novel strategies to prevent tumor angiogenesis can be developed, and form these, new anticancer therapies may arise.

• BMB Reports
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• 제53권1호
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• pp.56-63
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• 2020

The ubiquitin-proteasome system (UPS) and autophagy are two major degradative pathways of proteins in eukaryotic cells. As about 30% of newly synthesized proteins are known to be misfolded under normal cell conditions, the precise and timely operation of the UPS and autophagy to remove them as well as their tightly controlled regulation, is so important for proper cell function and survival. In the UPS, target proteins are labeled by small proteins called ubiquitin, which are then transported to the proteasome complex for degradation. Alternatively, many greatly damaged proteins are believed to be delivered to the lysosome for autophagic degradation. Although these autophagy and UPS pathways have not been considered to be directly related, many recent studies proposed their close link and dynamic interconversion. In this review, we'll focus on the several regulatory molecules that function in both UPS and autophagy and their crosstalk. Among the proposed multiple modulators, we will take a closer look at the so-called main connector of UPS-autophagy regulation, p62. Last, the functional role of p62 in the mitophagy and its implication for the pathogenesis of Parkinson's disease, one of the major neurodegenerative diseases, will be briefly reviewed.