• Environmental Analysis Health and Toxicology
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• v.19 no.3
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• pp.261-269
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• 2004

Although the biological functions of metallothioneins (MTs) are still being investigated, they have been suggested to be involved in detoxification of heavy metals, scavenging of free radicals, and protection against alkylating agents. MTs have been reported to be induced in most of animal tissues by heavy metals such as zinc, copper, mercury and cadmium, and the proteins have binding affinities to the metals. However, the presence or induction of MTs was reported not to be clear in leydig cells, which produce testosterone for the maturation of spermatozoa in male testes. In this study, we investigated the inducibility of metallothionein isomers by cadmium in cultured mouse leydig cells. Total RNA was extracted from the near confluent grown leydig cells and RT-PCR was Performed using the Primers which were synthesized on the basis of MT-1, 2, 3 and 4 cDNA from GenBank database. As results, MT-1 and MT-2 mRNA were found to be expressed in cadmium non-treated control cells and MT 1 mRNA expression was dose-dependent when leydig cells were treated with cadmium chloride. But MT-3 which is known to be brain specific and MT-4 which is another isoform of metallothionein, were not expressed. Other genes induced or depressed in cadmium treated leydig cells were also identified by microarray techniques.

• Molecules and Cells
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• v.37 no.10
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• pp.734-741
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• 2014

Histone modifications on major transcription factor target genes are one of the major regulatory mechanisms controlling adipogenesis. Plant homeodomain finger 2 (PHF2) is a Jumonji domain-containing protein and is known to demethylate the histone H3K9, a repressive gene marker. To better understand the function of PHF2 in adipocyte differentiation, we constructed stable PHF2 knock-down cells by using the mouse pre-adipocyte cell line 3T3-L1. When induced with adipogenic media, PHF2 knock-down cells showed reduced lipid accumulation compared to control cells. Differential expression using a cDNA microarray revealed significant reduction of metabolic pathway genes in the PHF2 knock-down cell line after differentiation. The reduced expression of major transcription factors and adipokines was confirmed with reverse transcription- quantitative polymerase chain reaction and Western blotting. We further performed co-immunoprecipitation analysis of PHF2 with four major adipogenic transcription factors, and we found that CCATT/enhancer binding protein (C/EBP)${\alpha}$ and C/EBP${\delta}$ physically interact with PHF2. In addition, PHF2 binding to target gene promoters was confirmed with a chromatin immunoprecipitation experiment. Finally, histone H3K9 methylation markers on the PHF2-binding sequences were increased in PHF2 knock-down cells after differentiation. Together, these results demonstrate that PHF2 histone demethylase controls adipogenic gene expression during differentiation.

• Horticultural Science & Technology
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• v.33 no.4
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• pp.575-584
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• 2015

Drought stress is a crucial environmental factor determining crop survival and productivity. The goal of this study was to clearly identify a new drought stress-tolerance gene in Brassica rapa. From KBGP-24K microarray data with the B. rapa ssp. pekinensis inbred line 'Chiifu' under drought stress treatment, a gene which was named BrDSR (B. rapa Drought Stress Resistance) was chosen among 738 drought-responsive unigenes. BrDSR function has yet to be determined, but its expression was induced over 6-fold by drought. To characterize BrDSR, the gene was isolated from B. rapa inbred line 'CT001' and found to contain a 438-bp open reading frame encoding a 145 amino acid protein. The full-length cDNA of BrDSR was used to construct an over-expression vector, 'pSL100'. Tobacco transformation was then conducted to analyze whether the BrDSR gene can increase drought tolerance in plants. The BrDSR expression level in T1 transgenic tobacco plants selected via PCR and DNA blot analyses was up to 2.6-fold higher than non-transgenic tobacco. Analysis of phenotype clearly showed that BrDSR-expressing tobacco plants exhibited more tolerance than wild type under 10 d drought stress. Taking all of these findings together, we expect that BrDSR functions effectively in plant growth and survival of drought stress conditions.

• Environmental Analysis Health and Toxicology
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• v.21 no.1 s.52
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• pp.71-79
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• 2006

Epidemiologic studies have showed a close correlation between arsenic exposure and heart disease such as, cardiovascular problem, ischemic heart disease, infarction, atherosclerosis and hypertension in human. It may increase the mortality of high risk group with heart disease. Regarding the mechanism studies of heart failure, blood vessel, vascular smooth muscle cells and endothelial cells have long been focused as the primary targets in arsenic exposure but there are only a few studies on the cardiomyocytes. In this study, the generation of oxidative stress by low dose of arsenic trioxide was investigated in rat cardiomyocytes. By direct measurement of reactive oxygen species and fluorescent microscopic observation using fluorescent dye 2',7'-dichlorofluorescin diacetate, reactive oxygen species were found to be generated without cell death, where cells are treated with 0.1 ppm arsenic for 24 hours. With the induction of reactive oxygen species, GSH level was decreased by the same treatment. However, DNA damage did not seem to be serious by DAPI staining, while high dose of arsenic (2 ppm for 24 hrs) caused fragmentation of DNA. To identify the molecular biomarkers of low-dose arsenic exposure, gene expression was also investigated with whole genome microarray. As results, 9,022 genes were up-regulated including heme oxygenase-l and glutathione S-transrerase, which are well-known biomarkers of oxidative stress. 9,404 genes were down-regulated including endothelial type gp 91-phox gene by the treatment of 0.1 ppm arsenic for 24 hours. This means that biological responses of cardiomyocytes may be altered by ROS induced by low level arsenic without cell death, and this alteration may be detected clearly by molecular biomarkers such as heme oxygenase-1.

• Korean Journal of Microbiology
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• v.51 no.1
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• pp.39-47
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• 2015

Secondary metabolism in actinomycetes has been known to be controlled by a small molecule, ${\gamma}$-butyrolactone autoregulator, the binding of which to each corresponding receptor leads to the regulation of the transcriptional expression of the secondary metabolites. We expected that expression of an autoregulator receptor or a pleiotropic regulator in a non-host was to be gained insight of effective production of new metabolic materials. In order to study the function of the receptor protein (seaR), which is isolated from Saccharopolyspora erythraea, we introduced the seaR gene to Streptomyces coelicolor A3(2) as host strains. An effective transformation procedure for S. coelicolor A3(2) was established based on transconjugation by Escherichia coli ET12567/pUZ8002 with a ${\varphi}C31$-derived integration vector, pSET152, which contained int, oriT, attP and $ermEp^*$ (erythromycin promotor). Therefore, the pEV615 was introduced into S. coelicolor A3(2) by conjugation and integrated at the attB locus in the chromosome of the recipients by the ${\varphi}C31$ integrase (int) function. Exconjugant of S. coelicolor A3(2) containing the seaR gene was confirmed by PCR and transcriptional expression of the seaR gene in the transformant was analyzed by RT-PCR. In case of S. coelicolor A3(2), a phenotype microarray was used to analyze the phenotype of transformant compared with wild type by seaR expression. After that, in order to confirm the accuracy of the results obtained from the phenotype microarray, an antimicrobial susceptibility test was carried out. This test indicated that sensitivity of the transformant was higher than wild type in tetracycline case. These results indicated that some biosynthesis genes or resistance genes for tetracycline biosynthesis in transformant might be repressed by seaR expression. Therefore, subsequent experiments, analysis of transcriptional pattern of genes for tetracycline production or resistance, are needed to confirm whether biosynthesis genes or resistance genes for tetracycline are repressed or not.

• Proceedings of the Korean Society for Bioinformatics Conference
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• 2006.02a
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• pp.83-89
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• 2006

Cellular functions are carried out by a concerted action of biochemical pathways whose components have genetic interactions. Abnormalities in the activity of the genes that constitute or modulate these pathways frequently have oncogenic implications. Therefore, identifying the upstream regulatory genes for major biochemical pathways and defining their roles in carcinogenesis can have important consequences in establishing an effective target-oriented antitumor strategy We have analyzed the gene expression profiles of human liver cancer samples using cDNA microarray chips enriched in liver and/or stomach-expressed cDNA elements, and identified groups of genes that can tell tumors from non-tumors or normal liver, or classify tumors according to clinical parameters such as tumor grade, age, and inflammation grade. We also set up a high-throughput cell-based assay system (cell chip) that can monitor the activity of major biochemical pathways through a reporter assay. Then, we applied the cell chip platform for the analysis of the HCC-associated genes discovered from transcriptome profiling, and found a number of cancer marker genes having a potential of modulating the activity of cancer-related biochemical pathways such as E2F, TCF, p53, Stat, Smad, AP-1, c-Myc, HIF and NF-kB. Some of these marker genes were previously blown to modulate these pathways, while most of the others not. Upon a fast-track phenotype analysis, a subset of the genes showed increased colony forming abilities in soft agar and altered cell morphology or adherence characteristics in the presence of purified matrix proteins. We are currently analyzing these selected marker genes in more detail for their effects on various biological Processes and for Possible clinical roles in liver cancer development.

• Journal of Acupuncture Research
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• v.22 no.3
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• pp.37-51
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• 2005

Mast cell is a cell that functions mainly in our body with a respect to inflammation and allergic response. Bee venom has been progressed in a study as a model related to mechanism in alleviation of pain until now, but it is being progressed in a study relevant to immunocyte in anti-inflammation or anti-allergic response. The present study is aimed to present the basis related to a future study of gene, by researching the influence of melittin and MCD Peptide, which are major ingredients in Bee venom, upon the expression of gene in the mast cell strain. In this study, it dealt with melittin and MCD Peptide respectively, in the effective concentration after passing though the experiment of cytotoxicity by using human mast cell strain. Also, with the respect in the aspect of expression in gene that changes at this time, information was obtained through the technique of analyzing microarray. Through experimental statistics, when regarding a case that global M is significant in more than 1 or -1, in melittin, all 7 genes were accelerated, and 8 inhibited. In MCDP, 7 genes were accelerated and 17 genes inhibited. The function in the body to which these genes are related, was associated with the protein binding within a cell, the activation in the function of lymphocyte, the acceptor related to macrophage antigen. In cell nucleus, substance related to GABA A receptor, protein associated with cAMP reactive element, substance related to complement system No.8 and to B-cell, protein substance related to polycystic kidney disease, substance related to inflammation, and the protein substance of influencing coagulation of blood. Through these results of analysis, it could obtain more useful materials in clarifying the mechanism of action in melittin and MCD peptide, which are in charge of mainly medical action in the abdomen. Also, it is thought that an in-depth study on the influence of main ingredients in Bee venom, the wholly honey bee venom aqua upon anti-allergic response or anti-inflammation are further required.

• Environmental Analysis Health and Toxicology
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• v.24 no.4
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• pp.321-332
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• 2009

본 연구의 목적은 다이아지논(Diazinon; O, O-diethyl O-[6-methyl-2 (1-methylethyl)-4-pyrimidinyl] phosphorothioate)에 노출된 모델 생물체(송사리)의 행동변화와 관련된 분자생물학적 기전 규명을 통하여 비정상적 행동의 모니터링을 위한 생물지표(biomarker)를 개발하는데 있다. 이를 위해 우선 suppression subtractive hybridization (SSH) 및 DNA microarray 기법을 활용하여 다양한 유전자를 스크리닝하였다. 다이아지논에 노출시킨 송사리에서 발현의 차이가 나는 상향 조절된 유전자 97개 (알려지지 않은 유전자 27개 포함)와 하향 조절된 유전자 99개 (알려지지 않은 유전자 60개 포함)를 동정 하였고 이들 중 이상행동과 관련되는 것으로 보이는 유전자 10개 (상향조절 5개, 하향조절 5개)를 선발하였다. 이들 중에서 primer 제작이 잘된 beta-1, Orla C3-1, parvalbumin 및 apolipoprotein E을 선발하여 그 유전자 발현을 real-time PCR 기법을 사용하여 정량적으로 모니터링 하였다. Orla C3-1, parvalbumin 및 apolipoprotein E는 고농도의 다이아지논 처리(1000 ppb; 24 h)에서 그 발현이 억제됨이 관찰되었다. 다이아지논 처리 시 신경질환 (알츠하이머 병 및 다운신드롬)에 관련된 apolipoprotein E와 근육세포의 유연화에 작용하는 parvalbumin 등의 발현억제는 송사리의 인지능력 교란 및 근육세포의 경직 등을 각각 유도하여 송사리의 비정상적 행동을 야기하는 것으로 판단되었다. 따라서 이들 생물지표는 신경독성물질에 의한 송사리 및 기타 어류의 이상행동의 변화의 감지에 활용될 수 있을 것으로 사료된다.

• Korean Journal of Biological Psychiatry
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• v.12 no.1
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• pp.42-61
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• 2005

Objectives:The ginsenoside Rg1 and Rb1, the major components of ginseng saponin, have neurotrophic and neuroprotective effects including promotion of neuronal survival and proliferation, facilitation of learning and memory, and protection from ischemic injury and apoptosis. In this study, to investigate the molecular basis of the effects of ginsenoside on neuron, we analyzed gene expression profiling of SH-SY5Y human neuroblastoma cells treated with ginsenoside Rg1 or Rb1. Methods:SH-SY5Y cells were cultured and treated in triplicate with ginsenoside Rg1 or Rb1($80{\mu}M$, $40{\mu}M$, $20{\mu}M$). The proliferation rates of SH-SY5Y cells were determined by MTT assay and microscopic examination. We used a high density cDNA microarray chip that contained 8K human genes to analyze the gene expression profiles in SH-SY5Y cells. We analyzed using the Significance Analysis of Microarray(SAM) method for identifying genes on a microarray with statistically significant changes in expression. Results:Treatment of SH-SY5Y cells with $80{\mu}M$ ginsenoside Rg1 or Rb1 for 36h showed maximal proliferation compared with other concentrations or control. The results of the microarray experiment yielded 96 genes were upregulated(${\geq}$3 fold) in Rg1 treated cells and 40 genes were up-regulated(${\geq}$2 fold) in Rb1 treated cells. Treatment with ginsenoside Rg1 for 36h induced the expression of some genes associated with protein biosynthesis, regulation of transcription or translation, cell proliferation and growth, neurogenesis and differentiation, regulation of cell cycle, energy transport and others. Genes associated with neurogenesis and neuronal differentiation such as SCG10 and MLP increased in ginsenoside Rg1 treated cells, but such changes did not occur in Rb1-group. Conclusion:Our data provide novel insights into the gene mechanisms involved in possible role for ginsenoside Rg1 or Rb1 in mediating neuronal proliferation or cell viability, which can elicit distinct patterns of gene expression in neuronal cell line. Ginsenoside Rg1 have more broad and strong effects than ginsenoside Rb1 in gene expression and related cellular physiology. In addition, we suggest that SCG10 gene, which is known to be expressed in neuronal differentiation during development and neuronal regeneration during adulthood, may have a role in enhancement of activity dependent synaptic plasticity or cytoskeletal regulation following treatment of ginsenoside Rg1. Further, ginsenoside Rg1 may have a possible role in regeneration of injured neuron, promotion of memory, and prevention from aging or neuronal degeneration.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.77.2-78
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• 2003

An EREBP/AP2-type transcription factor (CaPFl) was isolated by DDRT-PCR following inoculation of soybean pustule pathogen Xanthomonas axonopodis pv. glycines Bra which induces HR on pepper leaves. Genomic Southern blot analysis revealed that the CaPFl gene is present as a single copy within the hot pepper genome. The deduced amino acid sequence of CaPFl has two potential nuclear localization signals, a possible acidic activation domain, and an EREBP/AP2 motif that could bind to a conserved cis- element present in promoter region of many stress-induced genes. The mRNA level of CaPFl was induced by both biotic and abiotic stresses. We observed higher-level transcripts in resistance-induced pepper tissues than diseased tissues. Expression of CaPFl is also induced upon various abiotic stresses including ethephon, MeJA, cold stress, drought stress and salt stress treatments. To study the role of CPFI in plant, transgenic Arabidopsis and tobacco plants which express higher level of pepper CaPFl were generated. Global gene expression analysis of transgenic Arabidopsis by cDNA microarray indicated that expression of CaPFl in transgenic plants affect the expression of quite a few GCC box and DRE/CRT box-containing genes. Furthermore, the transgenic Arabidopsis and tobacco plant, expressing CaPFl showed tolerance against freezing temperature and enhanced resistance to Pseudomonas syrnigae pv. tabaci. Taken together, these results indicated that CaPFl is a novel EREBP/AP2 transcription factor in hot pepper plant and it may has a significant role(s) in regulation of biotic and abiotic stresses in plant.