• Molecules and Cells
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• 제42권1호
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• pp.17-27
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• 2019

Ubiquitin-specific protease 44 (USP44) has been implicated in tumor progression and metastasis across various tumors. However, the function of USP44 in prostate cancers and regulatory mechanism of histone-modifying enzymes by USP44 in tumors is not well-understood. Here, we found that enhancer of zeste homolog 2 (EZH2), a histone H3 lysine 27 methyltransferase, is regulated by USP44. We showed that EZH2 is a novel target of USP44 and that the protein stability of EZH2 is upregulated by USP44-mediated deubiquitination. In USP44 knockdown prostate cancer cells, the EZH2 protein level and its gene silencing activity were decreased. Furthermore, USP44 knockdown inhibited the tumorigenic characteristics and cancer stem cell-like behaviors of prostate cancer cells. Inhibition of tumorigenesis caused by USP44 knockdown was recovered by ectopic introduction of EZH2. Additionally, USP44 regulates the protein stability of oncogenic EZH2 mutants. Taken together, our results suggest that USP44 promotes the tumorigenesis of prostate cancer cells partly by stabilizing EZH2 and that USP44 is a viable therapeutic target for treating EZH2-dependent cancers.

• BMB Reports
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• 제55권12호
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• pp.595-601
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• 2022

Polycomb Repressive Complex 2 (PRC2) exhibits key roles in mammalian development through its temporospatial repression of gene expression. EZH1 or EZH2 is the catalytic subunit of PRC2 that mediates the mono-, di- and tri-methylation of histone H3 lysine 27 (H3K27me1/2/3), H3K27me2/me3 being a hallmark of facultative heterochromatin. PRC2 is a chromatin-modifying enzyme that is recruited to a limited number of "nucleation sites", spreads H3K27 methylation and fosters chromatin compaction. EZH1 and EZH2 exhibit differences in their expression patterns, levels of histone methyltransferase activity (HMT) in the context of PRC2, and DNA/nucleosome binding activity. This suggests that their roles in heterochromatin formation are disparate. Dysregulation of PRC2 activity leads to aberrant gene expression and is implicated in cancer and developmental diseases. In this review, we discuss the distinct function of PRC2/EZH1 and PRC2/EZH2 in the early and late developmental stages. We then discuss the cancers associated with PRC2/EZH1 and PRC2/EZH2.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.27-27
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• 2003

A diversified and concentrative approach of methylation player can be one of the most powerful studies in the understanding of global epigenetic modifications. Previous studies have suggested that DNA methylation contributes to transcriptional silencing through the several DNA methylation-mediated repression systems by hypermethylation, including methyltransferases (DNMTs), DNA methyltransferase association protein 1 (DMAPl), methyl-CpG binding domain (MBD), and histone deacetylases (HDACs). Assembly of these regulatory protein complexes act sequentially, reciprocally, and interdependently on the newly composed DNA strand through S phase. Therefore, these protein complexes have a role in coupling DNA replication to the designed turn-off system in genome. In this study, we attempted to address the role of DNA methylation by the functional analysis of the methyltransferase molecule, we described the involvement of DMAP1 and DNMTs in cell divistion and the effect of their loss. We also described distinct patterns that DMAP1 and DNMTs are spatially reorganized and displaced from condensing chromosomes as cells progress through mitosis in HeLa cell, COS7, and HIH3T3 cell cycle progressions. DNMT1, DNMT3b, and DMAP1 do not stably contact the genetic material during chromosome compaction and repressive expression. These finding show that the loss of activities of DNMTs and DMAP1 occure stage specifically during the cell cycle, may contribute to the integral balance of global DNA methylation. This is consistent with previous studies resulted in decreased histone acetyltransferases and HDACs, and differs from studies resulted in increased histone methyltransferases. Our results suggest that DNA methylation by DNMTs and DMAP1 during mitosis acts to antagonize hypermethylation by which this mark is epigenetical mitotic-specific methylation.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제36권4호
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• pp.243-249
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• 2010

Tissue engineered bone (TEB) can replace an autogenous bone graft requiring an secondary operation site as well as avoid complications like inflammation or infection from xenogenic or synthetic bone graft. Adult mesenchymal stem cells (MSC) for TEB are considered to have various ranges of differentiation capacity or multipotency by the donor site and age. This study examined the effect of age on proliferation capacity, differentiation capacity and bone morphogenetic protein-2 (BMP-2) responsiveness of human bone marrow stromal cells (hBMSC) according to the age. In addition, to evaluate the effect on enhancement for osteoblast differentiation, the hBMSC were treated with Trichostatin A (TSA) and 5-Azacitidine (5-AZC) which was HDAC inhibitors and methyltransferase inhibitors respectively affecting chromatin remodeling temporarily and reversibly. The young and old group of hBMSC obtained from the iliac crest from total 9 healthy patients, showed similar proliferation capacity. Cell surface markers such as CD34, CD45, CD90 and CD105 showed uniform expression regardless of age. However, the young group showed more prominent transdifferentiation capacity with adipogenic differentiation. The osteoblast differentiation capacity or BMP responsiveness was low and similar between young and old group. TSA and 5-AZC showed potential for enhancing the BMP effect on osteoblast differentiation by increasing the expression level of osteogenic master gene, such as DLX5, ALP. More study will be needed to determine the positive effect of the reversible function of HDAC inhibitors or methyltransferase inhibitors on enhancing the low osteoblast differentiation capacity of hBMSC.

• Current Research on Agriculture and Life Sciences
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• 제31권3호
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• pp.157-164
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• 2013

Chromatin remodelers that include histone methyl transferases (HMTases) are becoming a focal point in cancer drug development. The NSD family of three HMTases, NSD1, NSD2/MMSET/WHSC1, and NSD3/WHSC1L are bona fide oncogenes found aberrantly expressed in several cancers, suggesting their potential role for novel therapeutic strategies. Several histone modifiers including HMTase have clear roles in human carcinogenesis but the extent of their functions and regulations are not well understood, especially in pathological conditions. The extents of the NSDs biological roles in normal and pathological conditions remain unclear. In particular, the substrate specificity of the NSDs remains unsettled and discrepant data has been reported. NSD2/MMSET is a focal point for therapeutic interventions against multiple myeloma and especially for t(4;14) myeloma, which is associated with a significantly worse prognosis than other biological subgroups. Multiple myeloma is the second most common hematological malignancy in the United States, after non-Hodgkin lymphoma. Herein, as a first step before entering a pipeline for protein x-ray crystallography, we cloned, recombinantly expressed and purified the catalytic SET domain of NSD2. Next, we demonstrated the catalytic activities, in vitro, of the recombinantly expressed NSD2-SET on H3K36 and H4K20, its biological targets at the chromatin.

• Biomolecules & Therapeutics
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• 제17권1호
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• pp.12-16
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• 2009

One of the WHSC1/MMSET/NSD2 variant RE-IIBP is a histone H3-K27 methyltransferase with transcriptional repression activity. Overexpression of RE-IIBP in various types of leukemia suggests it's role in leukemogenesis. Here we identify two proteins, KRAB zinc finger protein ZNF 350 and enolase-1 as RE-IIBP interacting proteins by yeast two-hybrid screening and confirmed direct interaction in vivo and in vitro. Both proteins have been known for their role in transcriptional repression. Reporter assays using transient transfection demonstrated that both ZNF 350 and enolase-1 proteins synergistically repressed transcription with RE-IIBP, respectively. These results indicate both proteins have roles in RE-IIBP mediated transcriptional repression by involving co-repressor complex.

• Molecules and Cells
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• 제38권6호
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• pp.528-534
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• 2015

Hypoxia-inducible factor (HIF) is a key regulator of tumor growth and angiogenesis. Recent studies have shown that, BIX01294, a G9a histone methyltransferase (HMT)-specific inhibitor, induces apoptosis and inhibits the proliferation, migration, and invasion of cancer cells. However, not many studies have investigated whether inhibition of G9a HMT can modulate HIF-$1{\alpha}$ stability and angiogenesis. Here, we show that BIX01294 dose-dependently decreases levels of HIF-$1{\alpha}$ in HepG2 human hepatocellular carcinoma cells. The half-life of HIF-$1{\alpha}$, expression of proline hydroxylase 2 (PHD2), hydroxylated HIF-$1{\alpha}$ and von Hippel-Lindau protein (pVHL) under hypoxic conditions were decreased by BIX01294. The mRNA expression and secretion of vascular endothelial growth factor (VEGF) were also significantly reduced by BIX01294 under hypoxic conditions in HepG2 cells. BIX01294 remarkably decreased angiogenic activity induced by VEGF in vitro, ex vivo, and in vivo, as demonstrated by assays using human umbilical vein endothelial cells (HUVECs), mouse aortic rings, and chick chorioallantoic membranes (CAMs), respectively. Furthermore, BIX01294 suppressed VEGF-induced matrix metalloproteinase 2 (MMP2) activity and inhibited VEGF-induced phosphorylation of VEGF receptor 2 (VEGFR-2), focal adhesion kinase (FAK), and paxillin in HUVECs. In addition, BIX01294 inhibited VEGF-induced formation of actin cytoskeletal stress fibers. In conclusion, we demonstrated that BIX01294 inhibits HIF-$1{\alpha}$ stability and VEGF-induced angiogenesis through the VEGFR-2 signaling pathway and actin cytoskeletal remodeling, indicating a promising approach for developing novel therapeutics to stop tumor progression.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1998년도 Proceedings of UNESCO-internetwork Cooperative Regional Seminar and Workshop on Bioassay Guided Isolation of Bioactive Substances from Natural Products and Microbial Products
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• pp.136-136
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• 1998

L-asparaginyl and L- aspartyl residues in proteins are subject to spontaneous degradation reactions generating isomerized and racemized aspartyl derivatives. Proteins containing L-isoaspartyl and D-aspartyl residues usually have altered structures and diminished biological activities. These residues can be recognized and be repaired to normal L-aspartyl residues by protein L-isoaspartyl methyltransferase(PIMT), which is present at high levels in testis. Although testicular PIMT have been shown to be involved in either sperm motility or sperm maturation, it may play an important role in the repair of damaged sperm proteins during the prolonged period of epididymal transport and storage. In the present study, as a initial step toward elucidating the function of protein carboxylmethylation in testis, we purified PIMT from porcine testicular cytosol as a momeric 27,000 Da species by ammonium sulfate precipitation, DEAE-sephacel chromatography, SAH-liganded affinity chromatography, and gel filtration chromatography. The optimum pH for the reaction was 6.0. $K_{m}$ values of the enzyme for the S-adenosyl-L-methionine (SAM), synthetic oligopeptide(VYP-L-isoD-HA) and histone type II-As were 1.0 ${\mu}$M, 33.2 ${\mu}$M and 276 ${\mu}$M respectively. Consequently, properties of the porcine testicular PIMT is similar to that of other mammalian PIMTs.

• 농업과학연구
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• 제43권1호
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• pp.52-60
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• 2016

In animal reproduction, the quality of oocytes and embryos has been evaluated by the expression of specific molecules. Follistatin (FST), which was isolated from follicular fluid, binds and bio-neutralizes the TGF-${\beta}$ superfamily members. Previous studies using the bovine model showed FST could be an important molecular determinant of embryo developmental competence. However, the effect of FST treatment on porcine embryo developmental competence has not been established. In this study, the effect of exogenous FST on porcine embryo developmental competence was investigated during in vitro culture. FST (10 ng/ml) treatment induced a significant decrease in the rate of cell arrest at the 4-cell stage. The expression levels of DNA-methyltransferase 1 (DNMT1), histone deacetylase 1 (HDAC1), and histone deacetylase 2 (HDAC2) were decreased in 4-cell stage embryos. FST treatment also resulted in significant improvements in developmental competence of embryos in terms of blastocyst formation rate and OCT-4 mRNA levels, the latter being related to pluripotency. In conclusion, during in vitro culture, FST treatment significantly ameliorated 4-cell block during embryonic development and improved embryo developmental competence. Therefore, FST treatment may potentially have a functional role in porcine embryogenesis that is broadly applicable to enhance in vitro embryo development.

• Biomolecules & Therapeutics
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• 제27권2호
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• pp.231-239
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• 2019

Suppressor of Variegation 3-9 Homolog 2 (SUV39H2) methylates the lysine 9 residue of histone H3 and induces heterochromatin formation, resulting in transcriptional repression or silencing of target genes. SUV39H1 and SUV39H2 have a role in embryonic development, and SUV39H1 was shown to suppress cell cycle progression associated with Rb. However, the function of human SUV39H2 has not been extensively studied. We observed that forced expression of SUV39H2 decreased cell proliferation by inducing $G_1$ cell cycle arrest. In addition, SUV39H2 was degraded through the ubiquitin-proteasomal pathway. Using yeast two-hybrid screening to address the degradation mechanism and function of SUV39H2, we identified translationally controlled tumor protein (TCTP) as an SUV39H2-interacting molecule. Mapping of the interacting regions indicated that the N-terminal 60 amino acids (aa) of full-length SUV39H2 and the C-terminus of TCTP (120-172 aa) were critical for binding. The interaction of SUV39H2 and TCTP was further confirmed by co-immunoprecipitation and immunofluorescence staining for colocalization. Moreover, depletion of TCTP by RNAi led to up-regulation of SUV39H2 protein, while TCTP overexpression reduced SUV39H2 protein level. The half-life of SUV39H2 protein was significantly extended upon TCTP depletion. These results clearly indicate that TCTP negatively regulates the expression of SUV39H2 post-translationally. Furthermore, SUV39H2 induced apoptotic cell death in TCTP-knockdown cells. Taken together, we identified SUV39H2, as a novel target protein of TCTP and demonstrated that SUV39H2 regulates cell proliferation of lung cancer cells.