• Proceedings of the PSK Conference
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• 2001.04a
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• pp.48-55
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• 2001

When an elongating RNA polymerase encounters DNA damage on the template strand of a transcribed gene it can either be arrested by or be transcribed through the lesion. Lesions that arrest RNA polymerases are thought to be subject to transcription-coupled repair, whereas that damage that is bypassed can cause miscoding, resulting in mutations in the transcript (transcriptional mutagenesis). We have developed a technique using a plasmid-based luciferase reporter assay to determine the extent to which a particular type of DNA base modification is capable of causing transcriptional mutagenesis in vivo. The system uses Escherichia coli strains with different DNA repair backgrounds and is designed to detect phenotypic changes caused by transcriptional mutageneis under nongrowth conditions. In addition, this method is capable of indicating the extent to which a particular DNA repair enzyme (or pathway) suppresses the occurrence of transcriptional mutagenesis. Thus, this technique provides a tool with which the effects of various genes on non-replication-dependent pathways resulting in the generation of mutant proteins can be gauged.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2001.11a
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• pp.113-113
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• 2001

Stable MCF-7-ERE cells, in which pERE-Luc reporter gene has been stably integrated into the genome of the MCF-7 cells, were used to detect the estrogenic activity of various dietary flavonoids in either pure chemical or mixtures. Estradiol (E2) induced luciferase activity in dose dependent manner and this activity was inhibited by tamoxifen (Tam) concomitant treatment. A large series of flavonoids showed estrogenic activities, corresponding to EC5O values between 0.2 and 9 microM and their mixtures didn't show additive or synergistic effects. And we could find some structure and activity relationship. First, 4-methoxylation and catechol structure decreased estrogenic activities. Second, hydroxylation of 3 position reduced estrogenic effect. Third glycosides of flavonoids showed weak estrogenic activity or no activity. Interestingly, when tested at high concentrations, genistein, kaempferol, biochanin A and chrysin elicited luciferase induction higher than that of the maximum induction by estradiol. And these effects of genistein and kaempferol could not be fully inhibited with tamoxifen

• BMB Reports
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• v.42 no.3
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• pp.154-159
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• 2009

JARID1B (jumonji AT rich interactive domain 1B) is a large nuclear protein that is highly expressed in breast cancers and is proposed to function as a repressor of gene expression. In this paper, a phage display screen using the N-terminus of JARID1B as bait identified one of the JARID1B interacting proteins, namely PcG protein (Polycomb group) hPc2. We demonstrated that the C-terminal region, including the COOH box, was required for the interaction with the N-terminus of JARID1B. In a reporter assay system, co-expression of JARID1B with hPc2 significantly enhanced the transcriptional repression. These results support a role for hPc2 acting as a transcriptional co-repressor.

• BMB Reports
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• v.42 no.10
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• pp.691-696
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• 2009

The E2F gene family appears to regulate the proliferation and differentiation of events that are required for adipogenesis. Pref-1 is a transmembrane protein that inhibits adipocyte differentiation in 3T3-L1 cells. In this study, we found that the expression of pref-1 is regulated by the transcription factor E2F1. The expression of pref-1 and E2F1 was strongly induced in preadipocytes and at the late differentiation stage. Using luciferase reporter assay, ChIP assay and EMSA, we found that the -211/-194 region of the pref-1 promoter is essential for the binding of E2F1 as well as E2F1-dependent transcriptional activation. Knockdown of E2F1 reduced both pref-1 promoter activity and the level of pref-1 mRNA. Taken together, our data suggest that transcriptional activation of pref-1 is stimulated by E2F1 protein in adipocytes.

• Development and Reproduction
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• v.16 no.4
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• pp.379-384
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• 2012

Lhx8 (LIM homeobox 8) gene encodes a LIM homeodomain transcriptional regulator that is preferentially expressed in germ cells and critical for mammalian folliculogenesis. However, Lhx8 DNA binding sequences are not characterized yet. We aimed to identify and characterize a cis-acting sequence of germ-cell specific transcriptional factor, Lhx8. To identify Lhx8 DNA binding element, Cyclic Amplification of Sequence Target (CAST) Analysis was performed. Electrophoretic Mobility Shift Assay (EMSA) was processed for the binding specificity of Lhx8. Luciferase assay was for the transcriptional activity of Lhx8 through identified DNA binding site. We identified a putative cis-acting sequence, TGATTG as Lhx8 DNA binding element (LBE). In addition, Lhx8 binds to the LBE with high affinity and augments transcriptional activity of luciferase reporter driven by artificial promoter containing the Lhx8 binding element. These findings indicate that Lhx8 directly regulates the transcription of genes containing Lhx8 binding element in oocytes during early folliculogenesis.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2001.11a
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• pp.112-112
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• 2001

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmental toxin that activates the aryl hydrocarbon receptor (AhR) and disrupts multiple endocrine signaling pathways by enhancing ligand metabolism, altering hormone synthesis, down regulating receptor levels, and interfering with gene transcription. And TCDD-mediated gene transactivation via the AhR has been shown to be dependent upon estrogen receptor (ER) expression in human breast cancer cells. In the present study, we have examined the effect of natural estrogen, phytoestrognes and environmental estrogens on the regulation of CYP1A1 gene expression in MCF-7 human breast cancer cell line. that ER and AhR are co-expressed. pCYP1A1 -luc reporter gene was transiently transfected into MCF-7 cells. These cells were treated with various chemicals and then luciferase assay was carried out. 17be1a-estradiol significantly inhibited TCDD stimulated luciferase activity dose dependently and this inhibition was partially recovered by concomitant treatment of tamoxifen. 17beta-estradiol metabolites, 2-hydroxyestradiol and 16alpha-estriol resulted in less potent inhibitory effect than estradiol and synthetic estrogen, diethylstilbestrol (DES) showed no effect on CYP1A1 gene expression. This study demonstrated that estrogen down-regulated TCDD stimulated CYP1A1 expression via ER mediation. And we have found out that several flavonoids such as genistein, kaempferol, daidzein, naringenin, and alkylphenols such as nonylphenol, 4-octylphenol and resveratrol also inhibited TCDD induced CYP1A1 expression like estrogen.

• Journal of Korean Society of Forest Science
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• v.86 no.3
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• pp.261-269
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• 1997

Excised immature ovules and calli derived from the stems of cottonwood were bombarded with microprojectiles carrying plasmid DNA containing CaMV-35S promoter and ${\beta}$-glucuronidase(GUS) gene. After bombarded, the expression of GUS gene was detected by the assay of 5-bromo-4-chloro-3-indolyl-${\beta}$-gluconide(X-gluc). Transient gene expression was measured by counting the number of distinct regions of GUS activity per explant. As major parameters, the number of shots and the period of exposure to X-gluc after the bombardment were investigated for detecting GUS gene expression. In this experiment, the percents of GUS gene expression showing spots were 56.8 from immature ovules and 75.9 from micro-calli of cottonwood species. Among the treatments, two consecutive shots and 48 hour exposure produced about $25.75{\pm}2.77$(per ovule), $11.43{\pm}1.22$(per mini petridish) spots, respectively, Microprojectile particle bombardment provides a useful method to assay transient expression in both types of explants. Furthermore, our results represent that the excised ovule and/or the calli might be stably transformed by the biolistics.

• The Plant Pathology Journal
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• v.21 no.3
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• pp.288-292
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• 2005

Pathogenesis-related protein-1a (PR-1a) is strongly induced in tobacco plants by pathogen attack, exogenous salicylic acid (SA) application and by other developmental processes. In order to develop a rapid screening system for the selection of plant defense-inducing compounds originated from various sources, we have transformed tobacco Samsun NN plants with a chimeric construct consisting of GUS $(\beta-glucuronidase)$. In the $T_1$ generation, three transgenic lines having stable GUS expression were selected for further promoter analysis. Using GUS histochemical assay, we observed strong GUS induction driven by PR-1a promoter in PR1a-GUS transgenic tobacco leaves in response to the exogenous application of SA or benzol (1,2,3) thiadiazole-7-carbothioic acid S-methyl ester (BTH), a SA­derivative compound. In addition, GUS expression was maintained locally or systemically in PR1a-GUS transgenic line $\#5\;T_2$ generation) until after 3 days when they were treated with same chemicals. Our results suggested that the PR1a-GUS reporter gene system in tobacco plants may be applicable for the large-scale screening of defense-inducing substances.

• Food Science of Animal Resources
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• v.31 no.3
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• pp.398-404
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• 2011

TR4 has been suggested to play an important role in lipid metabolism in adipocytes. Although TR4 facilitates lipid accumulation during adipogenesis, the regulatory effect of TR4 on lipid storage in mature adipocytes remains unclear. We showed that TR4 inhibited the LXR agonist GW3965-mediated decrease of lipid accumulation in 3T3-L1 adipocytes. A reporter gene analysis revealed that TR4 suppressed LXR${\alpha}$ transcriptional activity, although LXR${\alpha}$ was unable to affect TR4 transcriptional activity. Moreover, adding TR4 resulted in reduced LXR${\alpha}$ binding to the LXR responsive element in a gel shift assay. Additionally, the suppressive effect of GW3965 on perilipin expression and lipid accumulation in 3T3-L1 adipocytes was abolished by TR4 overexpression. Taken together, our data demonstrate that TR4 plays an inhibitory role in LXR${\alpha}$-mediated suppression of lipid accumulation in 3T3-L1 adipocytes. This TR4 protective effect is mediated, in part, y blocking the suppressive effect of GW3965 on perilipin gene expression.

• Journal of The Korean Society of Grassland and Forage Science
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• v.34 no.3
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• pp.187-192
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• 2014

Alfalfa (Medicago sativa L.) is one of the most important forage legumes in the world. It has been demanded to establish the efficient transformation system in commercial varieties of alfalfa for forage molecular breeding and production of varieties possessing new characteristics. To approach this, genetic transformation techniques have been developed and modified. This work was performed to establish conditions for effective transformation of commercial alfalfa cultivars, Xinjiang Daye, ABT405, Vernal, Wintergreen and Alfagraze. GUS gene was used as a transgene and cotyledon and hypocotyl as a source of explants. Transformation efficiencies differed from 0 to 7.9% among alfalfa cultivars. Highest transformation efficiencies were observed in the cultivar Xinjiang Daye. The integration and expression of the transgenes in the transformed alfalfa plants was confirmed by polymerase chain reaction (PCR) and histochemical GUS assay. These data demonstrate highly efficient Agrobacterium transformation of diverse alfalfa cultivars Xinjiang Daye, which enables routine production of transgenic alfalfa plants.