H. pylori의 EGFR 신호전달에 미치는 영향 및 celecoxib의 위암발생 억제기전 연구

학위논문 (석사)-- 서울대학교 대학원 : 의학과, 2012. 8. 김나영.배경 : Helicobacter pylori (H. pylori)의 감염은 세포의 생존 조절능력을 저해하여 위암의 발생위험을 증가시킨다. H. pylori 감염은 세포내 EGFR의 활성화와 관련이 있으며, 하부단계의 phosphatidylinositol 3-OH kinase (PI3K)-Akt-Glycogen synthase kinase-3 (GSK3) 와 같은 경로를 활성화시켜 세포의 생존과 이동을 조절하는 것으로 알려져 있다. 또한 H. pylori감염은 COX-2의 과발현을 유발하는데, 이전 연구에서 COX-2 억제제인 celecoxib가 Akt 신호전달경로를 억제한다고 밝힌 바 있다. EGFR과 COX-2는 Akt 신호전달경로를 공유하며 긴밀한 영향을 주고받을 가능성이 있다. 이에 본 연구는 H. pylori가 EGFR 신호전달에 미치는 영향을 확인하고 celecoxib이 이를 억제하는지 확인하고자 하였다. 연구재료 및 방법 : AGS 위암세포주를 H. pylori (cagA+, vacA+) G27과 cagE 돌연변이를 가진 G27에 24시간동안 감염시킨 후 mRNA와 단백질의 발현을 확인하였다. COX-2, EGFR, TGF-ß, Snail, Slug, E-cadherin의 mRNA발현을 RT-PCR로확인하였다. 그리고 다양한 농도(0, 10, 20, 30 µmol/L)의 ceclecoxib를 처리하여 COX-2, EGFR, tAkt, pAkt와 pGSK3ß의 단백질 발현을 확인하였다. 결과 : AGS 위암세포주에서 wild type의 H. pylori 감염은 COX-2, EGFR, TGF-ß, Snail, Slug의 mRNA 발현을 증가시켰으며 E-cadherin의 mRNA발현을 감소시켰다. 또한 wild type의 H. pylori 감염은 COX-2, EGFR, pAkt, pGSK3ß 의 단백질 발현을 증가시킨 반면, cagE 돌연변이로 인해 IV형 분비계에 결함이 있는 H. pylori에 의한 감염은 EGFR을 활성화시키지 못하였다. Celecoxib은 H. pylori에 의해 과발현되는 COX-2 (p=0.015), EGFR (p=0.025), pAkt (p=0.025)와 pGSK3ß (p= 0.029)를 억제시켰다. 결론 : AGS 위암세포주에서IV형 분비계가 정상인H. pylori에 의한 감염은 EGFR 신호전달체계를 활성화시켰으며, celecoxib은 이를 억제하는 효과를 보였다.Background and aim: Helicobacter pylori (H. pylori) infection increases the risk of gastric cancer through disrupting the regulation of cell survival. H. pylori infection is associated with epithelial growth factor receptor (EGFR) activation. EGFR downstream targets, such as phosphatidyl inositol 3-OH kinase (PI3K)-Akt-glycogen synthase kinase-3 (GSK3) pathways, regulate cell survival and migration. H. pylori infection also induces cyclooxygenase-2 (COX-2) over-expression, and previous study suggested that selective COX-2 inhibitor, celecoxib, blocks Akt signaling pathways. COX-2 and EGFR may cross talk through Akt-signaling pathways. The aim of the present study was to evaluate the effect of H. pylori on EGFR signaling pathways and to find out whether celecoxib has inhibitory effect on the EGFR signaling pathway or not. Methods: AGS gastric epithelial cell lines were co-cultured with the toxigenic H. pylori cagA+, vacA+ G27 and the cagE- mutant of G27. The expressions of COX-2, EGFR, TGF-ß, Snail, Slug and E-cadherin were measured by real-time PCR. In the next, western blot analyses of COX-2, EGFR, total Akt (tAkt), phosphorylated Akt (pAkt) and pGSK3ß were carried out at various concentrations (0, 10, 20, 30μmol/L) of celecoxib treatment for 24 hours in H. pylori treated AGS cell lines. Results: H. pylori infection significantly up-regulated the mRNA levels of COX-2, EGFR,TGF-ß, Snail, Slug and down-regulated E-cadherin in RT-PCR. AGS cell lines treated with cagE- mutants, which have a defective type IV secretion system did not show EGFR up-regulation. Celecoxib had inhibitory effects on H. pylori-induced over-expression of COX-2 (p=0.015), EGFR (p=0.025), pAkt (p=0.025) and pGSK3ß (p=0.029) in AGS cell lines in Western blot analysis. Conclusion: Infection by H. pylori with intact type IV secretion system activates EGFR signal pathways in AGS cell lines and celecoxib has inhibitory effect on this pathway. These finding provide insights into the anti-gastric cancer effect of celecoxib.초록 I 목차 Ⅲ LIST OF TABLES Ⅳ LIST OF FIGURES Ⅴ 본문 서론 1 연구재료 및 방법 2 연구결과.. 5 고찰 13 참고문헌 17 국문초록 23Maste

중심립 유리과정에서 pericentrin 역할 연구

학위논문 (박사)-- 서울대학교 대학원 자연과학대학 생명과학부, 2017. 8. 이건수.A centrosome is composed of a centriole surrounded by protein matrix, called pericentriolar materials (PCM). The microtubule organization is the prime function of the centrosome. Cilia formation is another important function of the centrosome in quiescent cells. Duplication and segregation of centrioles occur in tight link to cell cycle. A daughter centriole is assembled next to the mother centriole during S phase, and remained in an engaged state until the cell exits M phase. New daughter centrioles may be generated only after the mother and daughter centrioles in the previous cycle are separated. Therefore, centriole separation is considered a licensing step for centriole duplication. However, it is largely unknown how centriole engagement is maintained and disrupted during the cell cycle. Pericentrin (PCNT) is a PCM protein which is important for maturation process of centrosome to become spindle poles during mitotic entry. PCNT is also involved in induction of centriole separation during mitotic exit. PCNT is specifically cleaved, which is considered an essential step for centriole separation during mitotic exit. The purpose of my research is to elucidate mechanistic aspects of PCNT functions in centriole engagement and separation during M phase. In chapter 1, I report that PCNT has to be phosphorylated by PLK1 in order to be a suitable substrate of separase. The phospho-resistant mutants of PCNT are not cleaved by separase and eventually inhibit centriole separation. Furthermore, phospho-mimetic PCNT mutants rescue centriole separation even in the presence of BI2536. Based on these results, I propose that PLK1 phosphorylation is a priming step for separase-mediated cleavage of PCNT and eventually for centriole separation. PLK1 phosphorylation of PCNT provides an additional layer of regulatory mechanism to ensure the fidelity of centriole separation during mitotic exit. In chapter 2, I generated PCNT knockout cell lines and analyzed the phenotypes in relation to PCM assembly and centriole association. Deletion of PCNT hardly affected interphase centrosomes but conferred defects in centrosome maturation in cells entering M phase. The centrioles in PCNT–deleted cells were prematurely separated in early phase of mitosis and frequently amplified in M phase-arrested cells. Abnormal multi-nuclear cells repeatedly appeared in PCNT-deleted cells at interphase. My results confirmed that PCNT is critical for centriole association during M phase.BACKGROUND 1 1. Structure of centrosome 1 1.1 Discovery of centrosome 1 1.2 Centrioles 1 1.3 Pericentriolar material (PCM) 2 2. Functions of centrosome 6 2.1 Microtubule network formation in interphase 6 2.2 Spindle formation during mitosis 6 2.3 Primary cilia formation in quiescent cells 7 3. Centrosome cycle 12 3.1 Initiation of centriole duplication 12 3.2 Centriole elongation 13 3.3 Centrosome maturation 13 3.4 Centrosome separation 14 3.5 Bipolar spindle formation 14 3.6 Centriole disengagement 14 4. Licensing mechanism for centriole duplication: Centriole engagement and disengagement 19 PURPOSE 23 CHAPTER 1. PLK1 regulation of PCNT cleavage ensures fidelity of centriole separation during mitotic exit 24 ABSTRACT 25 INTRODUCTION 26 MATERIAL AND METHODS 29 Plasmids, siRNA, and cell culture 29 Antibodies 30 Immunoprecipitation and immunoblot analyses 31 Immunostaining analysis 33 Statistical analysis 34 RESULTS 35 BI2536 blocks both PCM disassembly and centriole separation 35 PLK1 phosphorylation is necessary for PCNT cleavage 36 PLK1 phosphorylates PCNT in vivo 38 PCNT phosphorylation is necessary for centriole separation 40 PCNT and CEP215 are essential for centriole association 41 Dual functions of PLK1 phosphorylation of PCNT 43 Discussion 60 CHAPTER 2. Phenotypic analyses of PCNT-deleted cells 64 ABSTRACT 65 INTRODUCTION 66 MATERIAL AND METHODS 68 Plasmids, siRNA, and cell culture 68 Generation of knockout cell lines with CRISPR/CAS9 system 69 Antibodies 69 Immunoblot analysis 70 Immunostaining analysis 71 Statistical analysis 72 RESULTS 73 Generation of PCNTTP53 double knockout cell lines 73 Interphase centrosomes in the PCNT-deleted cells 73 Spindle poles in the PCNT-deleted mitotic cells 74 Active role of PCNT for centriole association 77 DISCUSSION 94 PERSPECTIVE 98 REFERENCES 101 ABSTRACT IN KOREAN (국문 초록) 108Docto

금나노 입자를 이용한 switchable-linker 기반 검출법의 응집과 침전반응에 영향을 미치는 요소에 관한 연구

학위논문 (석사)-- 서울대학교 대학원 : 농업생명과학대학 농생명공학부, 2018. 8. 최영진.Aggregation of gold nanoparticles (AuNPs) has been used in various fields for assorted purposes. In particular, detection of analytes through visual color changes induced by aggregation of AuNPs has been extensively studied in recent years. In our previous research, switchable-linker based detection system was introduced. In the system, sensing of specific targets is achieved based on the quantitative correlations between linkers, targets and particles. When the number of particles and linkers are ideally matched, a maximum aggregation occurs in suspension inducing aggregates settling. This linker range where aggregates precipitation dominantly occurs is called exhibiting visual color change (REVC). Since REVC plays a critical role in the system, it is important to gain an understanding of the factors that could affect in REVC formation. However, little research has been done about the basic mechanisms of REVC formation based on AuNPs aggregation and precipitation, especially focusing on the effects of particles size and surface area. In this study, the effects of total surface area of AuNPs in REVC formation were identified by comparing 3 different cases. In the first case, 11 nm and 19 nm-sized particles were used in the previous study. Under the same plasmon absorbance, as the size of particle increased, the total surface area decreased. In this case, REVC appeared at lower linker concentrations with a narrower range. After the addition of target streptavidin, REVC shifted more in 19 nm particles which had smaller surface area. As for the second case, 12, 18, and 24 nm-sized particles were used. In this case, the number of particles was identically adjusted, thus as the particle size increased, the total surface area of AuNPs increased. In larger surface area, REVC appeared at higher linker concentrations in a broader range which correspondsss to the previous results. With the addition of target streptavidin, REVC shifted less as the surface area increased. Lastly, the surface area of AuNPs was identically adjusted by controlling the number of particles using 13, 21, 30, and 36 nm-sized particles. Under the same surface area, REVC was formed at a similar linker concentrations regardless of individual particle sizes and REVC shifting occurred identically under the addition of target streptavidin. Three cases indicated that as the surface area of AuNPs decreased, REVC worked more sensitively as a sensor. As part of an effort to confirm the reaction time according to particles, the third case was intensively interpreted. When the surface area of AuNPs was identically controlled, it was observed that in 21 nm-sized particles, REVC was formed the slowest among others in observation time of 200 minutes. From DLS size measurements of aggregates and Stokes law, settling velocity of aggregates in REVC was calculated. At an early settling stage, aggregates of 30 and 36 nm-sized particles precipitated more rapidly. However, as time went by, settling speed of aggregates composed of 13 and 21 nm-sized particles reached beyond the speed of larger particles. At around 120 minutes, the settling velocity of aggregates composed of 13 nm-sized particles went over than that of 21 nm-sized particles. This phenomenon might be explained by the fact that smaller individual particles made clusters more rapidly in much bigger sizes, thus settling speed inversion seemed to be happened. The results of this study suggest the importance of the surface area in REVC formation and the aggregates precipitation speed inversion was revealed by the actual calculations using Stokes law. These can be contributed to the further development of switchable-linker based detection system and can be used as clues in other aggregation-based applications.Ⅰ. INTRODUCTION 10 Ⅱ. MATERIALS AND METHODS 13 2.1. Materials 13 2.2. Instrumentation 13 2.3. Preparation of gold nanoparticles 14 2.4. Preparation of streptavidin-coated gold nanoparticles (stAuNP) 18 2.5. Adjustment of the total surface area of AuNPs 22 2.6. Schematic explanations of the switchable-linker based detection system 24 Ⅲ. RESULTS AND DISCUSSION 28 3.1. Effects of AuNPs surface area on REVC formation under the different conditions 28 3.1.1. REVC formation under the same absorption spectra 28 3.1.2. REVC formation under the same number of particles 32 3.1.3. REVC formation under the same AuNPs surface area 35 3.2. Settling velocity of aggregates in REVC interpreted by the modified Stokes law 38 3.2.1. Aggregates sizes obtained by DLS and TEM measurements of 13, 21, 30, and 36 nm-sized particles 39 3.2.2. Settling velocity of aggregates in REVC using the modified Stokes law and occurrence of speed inversion phenomenon 47 Ⅳ. CONCLUSION 53 Ⅴ. REFERENCES 55 국문초록 59Maste

Cyclosporin A 치료의 부작용인 치은 과성장에 대한 azithromycin의 작용기전에 대한 연구

Dept. of Medical Science/석사[한글] 목적: Cyclosporin A (CsA)는 1970년대 이래로 장기이식이나 골수이식 수술을 받은 환자들에게 보편적으로 처방되는 면역억제제이다. 그러나 CsA는 획기적인 효과를 갖고 있음에도 불구하고, 신독성, 간독성 등 다양한 부작용을 나타내며, 치은과성장도 그 중 한가지이다. 이식 수술을 받은 후, CsA를 복용한 환자들에서 나타나는 치은과성장은 평균 30%로 높은 비율로 보고되었다. 최근 임상연구를 통해서 CsA로 인한 치은과성장은 macrolide 계열의 항생제인 azithromycin(AZI)에 의해 효과적으로 억제된다는 결과가 보고되었다. 그러나 현재까지 CsA에 의한 치은과성장의 유도기전과, AZI에 의한 억제기전은 명확히 밝혀지지 않았다. 그러므로, 본 연구에서는 CsA에 의한 치은과성장 유도 기전과 AZI에 의한 억제 기전을 밝히고자 한다. 방법: 치은 섬유아세포는 과거에 CsA를 복용하여 치은과성장을 보인 환자와 CsA를 복용한 경험이 없는 건강한 환자의 치은에서 explant culture로 분리하였다. 치은 섬유아세포의 CsA에 의한 증식률과 AZI에 의한 억제률은 MTT assay에 의해 측정하였고, 약물에 의한 치은 섬유아세포의 형태학적 변화를 광학 현미경을 이용하여 관찰하였다. 그리고 MMPs (matrix metalloproteinases)의 mRNA의 발현과 활성도는 각각 reverse transcription - polymerase chain reaction (RT-PCR)과 zymography로 조사하였다. 결과: CsA에 의한 세포의 증식률은 건강한 환자보다 CsA를 복용한 경험이 있는 환자로부터 분리한 치은 섬유아세포에서 뚜렷하게 증가되었다. 그러므로 이후의 연구는 CsA를 복용한 경험이 있는 환자의 치은 섬유아세포로 진행하였다. 120시간 동안 CsA (10ng/ml)와 AZI를 함께 처리한 경우, AZI의 농도에 비례하여 치은 섬유아세포의 증식률이 감소하였다. CsA에 의한 형태학적 변화는 관찰되지 않았지만, CsA와 AZI을 함께 처리했을 경우에는 타원형 모양으로 변화되었다. CsA를 처리한 치은 섬유아세포는 MMP-2의 mRNA 발현양과 활성도가 억제되었다. 그러나 AZI과 CsA를 함께 처리했을 경우에는 MMP-2의 mRNA 발현양은 CsA만 단독 처리한 경우와 비교하여 정상적으로 회복되었고, MMP-2와 MMP-9의 활성이 유도되었다. Membrane-type (MT)1 - MMP와 MT3 - MMP를 통해 latent MMP-2의 활성도를 억제한다고 보고된 testican 1은 CsA에 의해 mRNA의 발현이 많이 증가되었으나, AZI과 함께 처리한 치은 섬유아세포에서는 대조군과 비슷한 정도로 발현양이 감소되었다. 결론: AZI에 의한 세포증식 억제와 MMP-2의 활성도의 증가는 CsA에 의해 유도된 치은과성장을 억제하는데 중요하다. 핵심되는 말: cyclosporin A, azithromycin, 치은과성장, matrix metalloproteinase-2, testican 1 [영문]Cyclosporin A (CsA) is the most frequently used immunosuppressor in transplant surgery, but gingival overgrowth (GO) remains one of its adverse side effects. Many previous reports have demonstrated that GO can be effectively treated by azithromycin (AZI), a macrolide antibiotic of the azalide subclass that suppresses protein synthesis of both gram-positive and gram-negative aerobes. However, the mechanism by which AZI suppresses CsA-induced GO (CIGO) has not yet been elucidated. In the present study, we examined the inhibitory effect of AZI on CIGO and its mechanism of action, as well as the mechanism whereby CsA induces GO. Human gingival fibroblasts were isolated from the gingival tissues of healthy subjects and patients exhibiting CIGO. The cell proliferation was significantly increased by CsA exposure for 5 days in fibroblasts isolated from gingival tissue of patients taking CsA. In contrast, AZI significantly inhibited CsA-induced cell proliferation. Morphological alterations were observed in the gingival fibroblasts stimulated with CsA, AZI, or the combination of AZI and CsA. The mRNA level and activity of matrix metalloproteinase-2 (MMP-2) were inhibited by CsA, but were increased by the combination of AZI and CsA. The activities of the latent and active forms of MMP-2 were increased by AZI in a dose-dependent manner. Interestingly, both the latent and active forms of MMP-9 were secreted from gingival fibroblasts exposed to the combination of AZI and CsA. The increased mRNA expression level of testican 1 in CsA-treated gingival fibroblasts was inhibited by the combination of AZI and CsA in a dose-dependent manner. Testican 1 has been known to inhibit pro-MMP-2 activation by membrane-type 1 (MT1)-MMP or MT3-MMP. These results suggest that AZI alters the synthesis degradation of the extracellular matrix (ECM) by regulating the expression of testican 1, which consequently mediates the activation of MMP-2. Therefore, the induction of MMP-2 by AZI treatment might play an important role in the inhibitory mechanism of AZI on CIGO. Key words : cyclosporin A, azithromycin, gingival overgrowth, matrix metalloproteinase-2, testican 1ope

차세대 시퀀싱을 이용한 위내 세균과 위암 발병의 관련성에 관한 연구

학위논문 (박사)-- 서울대학교 대학원 : 의과대학 의학과 분자유전체전공, 2016. 2. 김나영.Introduction: Little is known about the role of gastric microbiota except for Helicobacter pylori (HP) in human health and disease. We compared the differences of human gastric microbiota according to gastric cancer or control and HP infection status and assessed the role of bacteria other than HP. Materials and methods: Gastric microbiota of 63 antral mucosa, 18 corpus mucosa samples were analyzed by barcoded 454-pyrosequencing of the 16S rRNA gene. Antral samples were divided into the four subgroups based on HP positivity in pyrosequencing and presence of cancer. The analysis was focused on bacteria other than HP, especially, nitrosating or nitrate reducing bacteria (NB). The changes of NB in antral mucosa of 16 subjects were followed-up. Results: The number of NB other than HP (non-HP-NB) was two times higher in the cancer groups than control groups but it did not reach statistical significance. The number of non-HP-NB tends to increase over time, but this phenomenon was prevented by HP eradication in the HP-positive control group, but not in the HP-positive cancer group. Discussion: We found out HP plays more important role than other bacteria in the gastric carcinogenesis.1. Introduction 1 2. Materials and Methods 3 3. Results 10 4. Discussion 30 5. References 36 국문초록 41Docto

PLK1에 의한 pericentrin 인산화가 중심립 유리현상에 미치는 영향 연구

학위논문 (석사)-- 서울대학교 대학원 : 생명과학부, 2013. 8. 이건수.중심체는 막이 없는 대표적인 세포소기관으로서, 세포분열기에 방추극으로 작용하여 방추사를 형성한다. 염색체 수 이상에 의해 발생되는 다극 방추체는 염색체불안정성 혹은 세포사멸을 일으킨다. 따라서, 정교한 중심체 수 조절기작은 세포가 자신의 유전물질을 안정적으로 딸세포에 전달해주는데 중요한 역할을 한다. 중심립 유리현상은 중심체의 수를 조절하는 핵심 기작으로 딸중심립이 모중심립에서 떨어지는 현상을 말한다. 또한 최근 연구결과에 따르면 separase에 의한 pericentrin의 절단이 중심립 유리현상을 일으키는 주요한 원인으로 밝혀졌다. 본 연구는 PLK1에 의해 조절되는 pericentrin 절단 기작에 대해 연구하였다. Pericentrin은 PLK1에 의해 2259번 및 2267번 아미노산이 인산화되며, 이 두 아미노산이 알라닌으로 치환된 돌연변이에서는 pericentrin의 절단 및 중심립 유리현상이 줄어듦을 관찰하였다. 이 결과는 PLK1이 중심립 유리현상을 pericentrin의 인산화를 통해 조절할 수 있음을 제안한다.Centrosome is a non-membrane bound organelle which functions as a spindle pole body during mitosis. Abnormalities in centrosome number cause multipolar spindles which result in chromosomal instability or cell death. Therefore the precise regulation of centrosome number is important for maintaining genome stability. During mitotic exit, procentrioles are disengaged from the mother centriole during mitotic exit, which is considered a licensing step for centriole duplication. It was recently reported that pericentrin cleavage is necessary and sufficient for centriole disengagement during mitosis. Here, I report that the pericentrin cleavage is regulated by PLK1. PLK1 phosphorylates S2259 and S2267 residues of pericentrin. FLAG-PCNT S2259A, S2267A in which these two residues are substituted to alanine is not cleaved during mitosis. Finally, centriole disengagement was inhibited in the pericentrin-depleted cells rescued with the phospho-resistant FLAG-PCNT mutants at S2259 and S2267. Based on the results, I propose that PLK1 phosphorylation is prerequisite to pericentrin cleavage and eventually to centriole disengagement during mitotic exit.ABSTRACT 1 CONTENTS 3 LIST OF FIGURES 5 INTRODUCTION 7 MATERIALS AND METHODS 9 DNA constructs and transfection 9 RNA interference 9 Cell culture and stable cell lines 10 Cell cycle synchronization and drug treatments 10 Antibodies 11 Immunofluorescence microscopy 11 Immunoblot analysis 12 RESULTS 13 The PLK1 activity is required for pericentrin cleavage during mitotic exit. 13 Specific phosphorylation of pericentrin is critical for its cleavage. 14 Phosphorylation at S2259 and S2267 residues is important for the pericentrin cleavage. 15 Specific phosphorylation of pericentrin is prerequisite to centriole disengagement during mitotic exit. 18 DISCUSSION 46 REFERENCES 48 ABSTRACT IN KOREAN(국문초록) 52Maste

소매여신에서의 손실분포 추정 방법 연구

학위논문(석사) --서울대학교 대학원 :통계학과,2007.Maste

Application of radon tracer to evaluate the interaction characteristics between groundwater and surface water with time variance around the groundwater heat pump system

학위논문 (석사)-- 서울대학교 대학원 : 지구환경과학부, 2015. 8. 이강근.The water exchange between groundwater and surface water has a significant impact on developing water resources. A geothermal energy system using indirect open loop ground source heat pump (GSHP) has been developed at Han River Environmental Research Center (HRERC) in Yangsu-ri, Yangpyeong-gun, Korea. For designing a high efficiency performance of the geothermal heat pump system in the study area, the characteristics of interaction between groundwater and surface water has to be figured out because the aquifer system of the study site is directly affected by Han Rivers. Therefore, radon-222 was used as an indicator because it has much higher concentration in groundwater than in surface water. In this study, 12 groundwater wells were used for monitoring radon concentration, hydrogeochemical parameters, and water level from May, 2014 to Apr., 2015. The statistical analysis and mixing analysis were also conducted for quantitative interpretation of the obtained radon and hydrogeochemical data. There are no significant correlations between the hydrogeochemical parameters and radon concentration. However, each group classified by the cluster analysis was reflected its own hydrogeochemical properties. The spatial distribution of radon concentration was affected by seasonal effect and fluctuation of Han River stage from surrounding dam activity. The results of mixing analysis represented the quantitative interpretation of radon data. This study demonstrated radon-222 can be used as an appropriate environmental tracer in examining the characteristics of interaction between groundwater and surface water in consideration of river discharge rate. These results help to provide useful background information on the development of GSHP in the riverside area.ABSTRACT ……………………………………………ⅰ TABLE OF CONTENTS …………………………… ⅲ FIGURE LIST ……………………………………………ⅴ TABLE LIST …………………………………………… ⅷ 1. INTRODUCTION ……………………………………… 1 1.1 Research Backgrounds ………………………… 1 1.2 Objectives and Scope …………………………… 4 2. LITERATURE REVIEW ……………………………… 5 2.1 Radon as a Tracer ……………………………… 5 3. STUDY AREA ………………………………………… 7 3.1 Site Description …………………………………… 7 3.2 Geological Characteristics ………………………… 9 3.3 Hydrological Characteristics ……………………… 18 4. METHODS AND MATERIALS ……………………… 21 4.1 Well Installation …………………………………… 21 4.2 Water Sampling and Hydrogeochemical Analysis 24 4.3 Monitoring of Groundwater Flow ………………… 25 4.4 Measurement of Radon Concentration …………… 26 4.5 The Mass Balance Equation for Mixing Analysis 28 4.6 Statistical Analysis of Data ……………………… 31 5. RESULTS AND DISCUSSION ……………………… 33 5.1 Long Term Monitoring of Groundwater and Surface Water ……………………………………………………… 33 5.2 Statistical Application to Interpret Hydrogeochemical Properties in Groundwater ……… 45 5.3 Short Term Monitoring of Groundwater and Surface Water …………………………………………… 54 5.4 Mixing Analysis by Using Radon-222 ………… 58 6. SUMMARY AND CONCLUSTION ………………… 69 REFERENCE …………………………………………… 72Maste

에스트로겐 수용체-알파의 Ufmylation이 에스트로겐 신호전달과정에 미치는 영향에 관한 연구

학위논문 (석사)-- 서울대학교 대학원 : 생명과학부, 2014. 2. 정진하.UFM1, ubiquitin-like protein, is post-translationally conjugated to cellular proteins. The ufmylation reaction is catalyzed by an UFM1-activating E1 enzyme (UBA5), an UFM1-conjugating E2 enzyme (UBC1), and UFM1 E3 ligases (UFL1), in a manner similar to ubiquitination. Estrogen receptor α(ERα) is a member of a large conserved superfamily of steroid hormone nuclear receptors, acting as ligand-regulated transcription factor. ERα regulates many physiological pathways in response to its ligand, E2 (17β-estradiol). ERα is undergoing different types of post-translational modifications, such as phosphorylation, sumoylation, acetylation, and ubiquitination, which regulate its transcriptional activation and/or stability. Here, ERα was found to be a target for modification by UFM1. I also identified the major UFM1 acceptor site is in the AF1 domain. Moreover, knock down of UBA5 by RNA interference reduced the ERα transcriptional activity and accelerated E2-induced proteolysis. These results indicate that UFM1 modification plays a critical role in positive regulation of ERα function in transcriptional activation by preventing its degradationAbstract 1 Introduction 2 Materials and Methods 6 Plasmids and antibodies Cell culture and transfections Assays for ufmylation and deufmylation Ni-NTA-agarose pull-down analysis and immunoprecipitation Luciferase assays Results 9 ERα is a substrate for UFM1 modification Formation of poly-UFM1 chain via K69-linked isopeptide bonds Identification of the UFM1 acceptor sites in ERα Reversal of ERα ufmylation by UFSP2 Requirement of ERα ufmylation for E2-induced ERα transactivation Effect of ERα ufmylation for E2-induced ERα proteolysis Inverse correlation between E2-induced ufmylation and ubiquitination of ERα Discussion 31 References 36 국 문 초 록 40Maste