• The Korean Journal of Physiology and Pharmacology
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• 제20권4호
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• pp.367-378
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• 2016

Recently, it was reported that the role of mitochondria-reactive oxygen species (ROS) generating pathway in cisplatin-induced apoptosis is remarkable. Since a variety of molecules are involved in the pathway, a comprehensive approach to delineate the biological interactions of the molecules is required. However, quantitative modeling of the mitochondria-ROS generating pathway based on experiment and systemic analysis using the model have not been attempted so far. Thus, we conducted experiments to measure the concentration changes of critical molecules associated with mitochondrial apoptosis in both human mesothelioma H2052 and their ${\rho}^0$ cells lacking mitochondrial DNA (mtDNA). Based on the experiments, a novel mathematical model that can represent the essential dynamics of the mitochondrial apoptotic pathway induced by cisplatin was developed. The kinetic parameter values of the mathematical model were estimated from the experimental data. Then, we have investigated the dynamical properties of this model and predicted the apoptosis levels for various concentrations of cisplatin beyond the range of experiments. From parametric perturbation analysis, we further found that apoptosis will reach its saturation level beyond a certain critical cisplatin concentration.

• Bulletin of the Korean Chemical Society
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• 제30권2호
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• pp.323-326
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• 2009

SR proteins are essential splicing regulators and also modulate alternative splicing events, which function both as redundant and substrate-specific manner. The Drosophila B52/SRp55, a member of the SR protein family, is essential for the fly development in vivo, as deletion of B52 gene results in lethality of animals at the second instar larval stage. Identification of the splicing target genes of B52 thus should be crucial for the understanding of the specific developmental role of B52 in vivo. In this study, we performed whole-genome DNA microarray experiments with a B52- knock-out animal. Analysis of the microarray data not only provided the B52-dependent gene expression signature, but also revealed a larval-stage specific, alternative splicing target gene of B52. Our result thus provides a starting point to understand the essential function of B52 at the organismal level.

• Journal of Microbiology and Biotechnology
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• 제9권6호
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• pp.881-884
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• 1999

The gene encoding the RNA-dependent RNA polymerase of the hepatitis C virus was cloned and expressed with a C-terminal hexahistidine tag. The protein was purified from Escherichia coli to near homogeneity and characterized in vitro. When the 21 amino acids from the C-terminus of the protein were deleted, an inclusion body was not formed and a better purification yield was achieved. However, the activity of the purified enzyme decreased compared to that of the full length protein. The purified enzyme did exhibit ribonucleotide-incorporation activity on an in vitro transcribed RNA containing the 3' end of the HCV genome. It also possessed ribonucleotide incorporation activity, to a lesser extent, on in vitro transcribed foreign RNA templates when RNA or DNA primers were present. The activity was higher with DNA primers than with RNA primers. Accordingly, this assay system will facilitate the screening of inhibitors for hepatitis C virus replication.

• BMB Reports
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• 제54권8호
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• pp.413-418
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• 2021

ErbB3-binding protein 1 (EBP1) is a multifunctional protein associated with neural development. Loss of Ebp1 leads to upregulation of the gene silencing unit suppressor of variegation 3-9 homolog 1 (Suv39H1)/DNA (cytosine 5)-methyltransferase (DNMT1). EBP1 directly binds to the promoter region of DNMT1, repressing DNA methylation, and hence, promoting neural development. In the current study, we showed that EBP1 suppresses histone methyltransferase activity of Suv39H1 by promoting ubiquitin-proteasome system (UPS)-dependent degradation of Suv39H1. In addition, we showed that EBP1 directly interacts with Suv39H1, and this interaction is required for recruiting the E3 ligase MDM2 for Suv39H1 degradation. Thus, our findings suggest that EBP1 regulates UPS-dependent degradation of Suv39H1 to govern proper heterochromatin assembly during neural development.

• 대한의생명과학회지
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• 제29권4호
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• pp.280-286
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• 2023

In humic substances, fulvic acid (FA) is a subclass of diverse compounds known as humic substances, which are by-products of organic degradation from microorganisms. FA can suppress the proliferation of tumor cells. Despite numerous studies, the exact mechanism for the various effects of FA is not clearly understood. Based on results demonstrating anti-proliferation effects on human cancer, we investigated whether FA has similar effects on lung cancer in this study. Firstly, the anti-cancer effect of FA in pulmonary epithelial tumor cell lines (TC-1 cells) was examined by confirming its inhibitory effect on the cell proliferation of TC-1 cells. TC-1 cell proliferation was reduced by FA on a dose-dependent and time-dependent manner. After 24 hours of FA treatment, cell morphological changes such as cell volume decrease, non-adherence and increased number of apoptotic cells were clearly observed. In addition, FA induced a DNA ladder pattern by increased of DNA fragments in TC-1 cells. In the intracellular regulatory pathway by FA, we confirmed that FA induced the reduction of the anti-apoptotic protein, Bcl-2 protein levels. These results indicate that FA has anticancer effect by inducing intracellular apoptotic pathway. Further research on the mechanism of anticancer effects will be basic data for the development of potential anticancer drugs.

• Journal of Photoscience
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• 제9권2호
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• pp.482-484
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• 2002

UV irradiation activates various intracellular signaling pathways causing cell death in a DNA damage-dependent and an independent manner. As DNA photoproducts, major forms of DNA damage, are maximally formed by UV light at 260-nm, short wavelength UV (UVC) is more harmful than middle wavelength UV (UVB). However, the differences or similarities in responses of DNA damage-independent intracellular signaling molecules to UVB and UVC are not elucidated. We examined activation of signaling molecules towards apoptosis in normal human fibroblastic cells after irradiation with UVB or UVC at a dose generating the equal amount of DNA photoproducts. Both UVB and UVC induced transient phosphorylation of ERK and sustained phosphorylation of p38. Phosphorylation of p53 at Ser15 and at Ser392 residues were also observed, which were inhibited by a phosphoinositide 3-kinase inhibitor, wortmannin. In contrast, an antioxidant N-acetyl-cysteine and a p38 inhibitor SB203580 suppressed only Ser392 phosphorylation, suggesting that UV-induced oxidative stress and p38 activation were involved in the phosphorylation of this site. The apoptic signals such as mitochondrial cytochrome C release and annexin V binding were then observed. Overall, no difference was found in chronological responses of p53, MAPK, and apoptosis between UVB-irradiated and UVC-irradiated cells. These results suggested that DNA damage-independent intracellular signaling molecules similarly responded to UVB and UVC when the equal level of DNA photoproducts were generated.

• Animal cells and systems
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• 제6권3호
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• pp.263-269
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• 2002

The p53 tumor suppressor gene is among the most frequently mutated and studied genes in human cancer, but the mechanisms by which it sur presses tumor formation remain unclear. DNA damage regulates both the protein levels of p53 and its affinity for specific DNA sequences. Stabilization of p53 in response to DNA damage is caused by its dissociation from Mdm2, a downstream target gene of p53 and a protein that targets p53 for degradation in the proteosome. Recent studies have suggested that phosphorylation of human p53 at Ser20 is important for stabilizing p53 in response to DNA damage through disruption of the interaction between Mdm2 and p53. We generated mice with an allele encoding changes at Ser20, known to be essential for p53 accumulation following DNA damage, to enable analyses of p53 stabilization in vivo. Our data showed that the mutant p53 was clearly defective for full stabilization of p53 in response to DNA damage. We concluded that Ser20 phosphorylation is critical for modulating the negative regulation of p53 by Mdm2, probably through phosphorylation-dependent inhibition of p53-Mdm2 interaction in the physiological context.

• 미생물학회지
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• 제34권3호
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• pp.132-136
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• 1998

자연환경의 미생물군집을 분자생태학적 방법으로 분석하기 위해서는 오염되지 않은 순수한 핵산의 분리가 필요하다. 해양퇴적물에서 핵산을 추출할 때 같이 추출되는 부식물질(humic substances)은 핵산중폭반응을 저해하기 때문에, 이를 제거하기 위한 네 가지의 방법, 아가로즈젤 전기영동후 용출, Sephadex G-75 젤, Hydroxyapatite 젤, 그리고 PVPP(poluvinylpolypyrrolidone) microspin column 방법에 대해 그 효율성을 비교하였다. 조산된 방법 중에서 PVPP microspin column을 이용하면 건조 퇴적물 1g당 $4.8{\mu}g$의 순수한 DNA를 신속하고 간편하게 얻을 수 있었고, 이 방법으로 분리된 DNA를 PCR의 주형으로 사용한 결과 16S rRNA 유전자의 거의 전 범위에 해당하는 1.5 kb 크기의 PCR 산물을 얻을 수 있었다.

• 한국수정란이식학회지
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• 제25권2호
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• pp.111-116
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• 2010

Sperm chromatin integrity is essential for successful fertilization and development of an embryo. Reported here is a quantification of DNA fragments which is intimately associated with reproductive potential to provide one of criteria for sperm chromatin integrity. Three sperm populations were considered: CONTROL (no treatment), UV irradiation (48mW/$cm^2$, 1h) and $H_2O_2$ (oxidative stress induced by hydrogen peroxide, 10 mM, 50 mM and 100 mM). DNA fragments in boar sperm were evaluated by using ligation-mediated quantitative real-time polymerase chain reaction (LM-qPCR) assay, which relies on real-time qPCR to provide a measure of blunt 5' phosphorylated double strand breaks in genomic DNA. The results in agarose gel electrophoresis showed no significant DNA fragmentation and no dose-dependent response to $H_2O_2$. However, the remarkable difference in shape and position was observed in melting curve of LM-qPCR. This result supported that the melting curve analysis of LM-qPCR presented here, could be more sensitive and accurate than previous DNA fragmentation assay method.

• 대한방사선치료학회지
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• 제4권1호
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• pp.71-77
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• 1990

본 연구는 악하선 세포를 배양하고 전리 방사선을 조사하여 배양 세포의 DNA 합성능의 변화와 염색체의 구조적 이상을 연구하였다. SG세포는 DME배지에 $10\%$ FBS와 항생제, fungizone등이 첨가된 배양액에 배양하였다. 배양 악하선 세포에 대한 전리 방사선의 조사는 선량별로 $^{60}Co$ gamma선원을 이용하여 (dose rate 58.4 rad/min)실시하였다. 배양세포의 DNA 합성에 관한 방사선의 효과는 $^{3}H-TdR$의 동조율(incorporation)을 측정하여 평가하였다. 전리 방사선에 의하여 유발된 배양 악하선 세포의 염색체 이상을 관찰하기 위한 염색체 표본제작은 일반적으로 널리 사용되고 있는 방법에 따랐으며, 제작된 슬라이드는 Giemsa염색액으로 단염색 하였다. 본 실험에서 얻어진 결과를 요약하면 다음과 같다. 1. 배양 악하선 세포의 DNA 합성능은 전리 방사선의 양이 증가함에 따라 그 합성능이 감소하였다. 2. 방사선 조사후 배양2일째에 그 DNA 합성능이 회복되었다. 3. 본 연구에 나타난 염색체 이상은 염색체 절단(single break과 double break), 결손, triradius등이었으며 polyploid도 관찰되었다. 4. 전리 방사선에 의해 유발된 염색체 이상은 선량의 증가에 따라 그 발생빈도 역시 증가되었다.