• International Journal of Industrial Entomology and Biomaterials
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• v.7 no.2
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• pp.181-189
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• 2003

We describe here the complete nucleotide sequence and the exon-intron structure of the luciferase gene of the firefly, Pyrocoelia rufa. The luciferase gene of the P. rufa firefly consisted of six introns and seven exons coding for 548 amino acid residues. From the translational start site to the end of last exon, however, the genomic DNA length of the P. rufa luciferase gene from the Korean and Chinese samples spans 1,968 bp and 1983 bp, respectively, and 3 amino acid residues were different to each other. Additionally, we also analyzed mitochondrial cytochrome oxidase I(COI) gene of the Chinese P. rufa fireflies. Analysis of DNA sequences from the mitochondrial COI protein-coding gene revealed 4 mitochondrial DNA sequence-based haplotypes with a maximum divergence of 0.7％. With the 20 P. rufa haplotypes found in Korea, phylogenetic analyses using PAUP and PHYLIP subdivided the P. rufa into three clades, termed clades A and B for the Korean sample, and clade C for the Chinese sample.

• Preventive Nutrition and Food Science
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• v.8 no.1
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• pp.61-65
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• 2003

This study was investigated to identify the regulatory mechanism of ACC gene expression by hormones and nutrition. The fragment of ACC promoter I (PI) -220 bp region was recombined to pGL3-Basic vector with luciferase as a reporter gene. The primary hepatocyte from the rat was used to investigate the regulation of ACC PI activity. ACC PI (-220 bp)/luciferase chimeric plasmid was transfected into primary rat hepatocyte by using lipofectin. ACC PI activity was shown by measuring luciferase activity. The addition of insulin, dexamethasone, and triiodothyronine to the culture medium increased the activity of ACC PI by 2.5-, 2.3- and 1.8-fold, respectively. In the presence of 1 $\mu$M dexamethasone, the effects of insulin was amplified about 1.2-fold showing the additional effects of dexamethasone. Moreover the activity of luciferase was increased by insulin, dexamethasone, and triiodothyronine treatment approximately 4-fold. These results indicated that insulin, dexamethasone and thyroid hormone coordinately regulate ACC gene expression via regulation of promoter I activity. On the -220 to ＋21 region of ACC PI, the addition of the glucose to the culture medium increased the activity of ACC PI. With 25 mM glucose, luciferase activity increased by 7-fold. On the other hand, on the -220 bp region, ACC PI activity was not changed by polyunsaturated fatty acids. Therefore, it can be postulated that there are response elements for insulin, triiodothyronine, dexamethasone, and glucose, but not PUFAs on the -220 bp region of ACC PI.

• The Korean Journal of Food And Nutrition
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• v.30 no.5
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• pp.900-907
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• 2017

This study was performed to investigate the body fat-lowering effect of garlic powder in peroxisome proliferator-activated receptor ${\gamma}$ coactivator-$1{\alpha}$(PGC-$1{\alpha}$)-luciferase transgenic mice (TG). In this study, we generated transgenic mice with a PGC-$1{\alpha}$ promoter (-970/+412 bp) containing luciferase as a reporter gene. Mice were fed a 45% high-fat diet for 8 weeks to induce obesity. Subsequently, mice were maintained on either a high-fat control diet (CON), or high-fat diets supplemented with 2% (GP2) or 5% (GP5) garlic powder for an additional 8 weeks. Dietary garlic powder reduced the body weight in the GP2 and GP5 groups, compared to the CON group. Furthermore, garlic supplementation significantly decreased the plasma levels of triglycerides, total cholesterol, and leptin in the GP5 group, compared to the CON group. Specifically, luciferase activity in liver, white adipose tissue (WAT), and brown adipose tissue (BAT) was increased by garlic supplementation in a dose-dependent manner. These results suggest that the body fat-lowering effect of garlic powder might be related to PGC-$1{\alpha}$ by the increase in luciferase activity in liver, WAT, and BAT. Furthermore, transgenic mice might be useful for evaluating the body fat-lowering effect of various health functional foods.

• Korean Journal of Agricultural Science
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• v.48 no.2
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• pp.353-365
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• 2021

Previously, we reported that tissue factor (Tf) was included in the list of differentially expressed genes as an upregulated gene in a rejected porcine heart after xenotransplantation into monkey. In this study, we analyzed that expression of Tf in aortic endothelial cells (pAEC) isolated from alpha 1,3-galactosyltransferase knockout pig in response to allogeneic porcine serum and xenogeneic human serum. The consequence was significant upregulation of Tf expression by responding to human serum compared with porcine serum. To analyze the function of Tf gene as a promoter, we constructed reporter vectors for expression of luciferase linked to 1,246 and 787 base pairs of porcine Tf (pTF1246 and pTF787), and 535 base pairs of human TF (hTF535) sequences including putative promoter regions and AP-1 biding site at the 5' end. The reporter vectors were transfected into pAEC including cytomegalovirus enhancer/chicken β-actin (CAG)-luciferase vector as a control. Luciferase assay showed that all of the promoters were insufficient to express luciferase compared with CAG promoter in basic culture conditions. Notably, pTF1246, pTF787, and hTF535 led to a significant increase of luciferase expression in response to human serum compared with porcine serum while no change of CAG. pTF1246 and pTF787 showed higher expression than hTF535. Taken together, our findings suggest that pTF1246 and pTF787 promoters could mediate target gene expression specifically at xenogeneic stress condition.

• Journal of Ginseng Research
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• v.46 no.2
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• pp.248-254
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• 2022

Background: Zinc homeostasis is essential for human health and is regulated by several zinc transporters including ZIP and ZnT. ZIP4 is expressed in the small intestine and is important for zinc absorption from the diet. We investigated in the present study the effects of Panax ginseng (P. ginseng) extract on modulating Zip4 expression and cellular zinc levels in mouse Hepa cells. Methods: Hepa cells were transfected with a luciferase reporter plasmid that contains metal-responsive elements, incubated with P. ginseng extract, and luciferase activity was measured. Using ⁶⁵ZnCl₂, zinc uptake in P. ginseng-treated cells was measured. The expression of Zip4 mRNA and protein in Hepa cells was also investigated. Finally, using a luciferase reporter assay system, the effects of several ginsenosides were monitored. Results: The luciferase activity in cells incubated with P. ginseng extract was significantly higher than that of control cells cultured in normal medium. Hepa cells treated with P. ginseng extract exhibited higher zinc uptake. P. ginseng extract induced Zip4 mRNA expression, which resulted in an enhancement of Zip4 protein expression. Furthermore, some ginsenosides, such as ginsenoside Rc and Re, enhanced luciferase activity driven by intracellular zinc levels. Conclusion: P. ginseng extract induced Zip4 expression at the mRNA and protein level and resulted in higher zinc uptake in Hepa cells. Some ginsenosides facilitated zinc influx. On the basis of these results, we suggest a novel effect of P. ginseng on Zip4-mediated zinc influx, which may provide a new strategy for preventing zinc deficiency.

• Journal of Life Science
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• v.23 no.2
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• pp.167-174
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• 2013

Peroxiredoxin 6 (Prx 6) is a member of the thiol-specific antioxidant protein family, which may play a role in protection against oxidative stress and in regulating phospholipid turnover. The aim of this study was to determine whether a human Prx 6/Luc vector was stably expressed and responded to antioxidants in a lung cell line (NCI-H460). To achieve this, the luciferase signal, hPrx 6 mRNA expression, and superoxide dismutase (SOD) activity were measured in transfectants with a hPrx 6/Luc plasmid after treatment with four antioxidant extracts, including Korea white ginseng (KWG), Korea red ginseng (KRG), Liriope platyphylla (LP), and red Liriope platyphylla (RLP). First, the hPrx 6/Luc plasmid was successfully constructed with DNA fragments of human Prx 6 promoter, amplified by PCR using genomic DNA isolated from NCI-H460 cells, and cloned into the pTransLucent reporter vector. The orientation and sequencing of the hPrx 6/Luc plasmid were identified with restriction enzyme and automatic sequencing. A luciferase assay revealed significant enhancement of luciferase activity in the four treatment groups compared with a vehicle-treated group, although the ratio of the increase was different within each group. The KRG- and LP-treated groups showed higher activity than the KWG- and RLP-treated groups. Furthermore, the luciferase activity against RLP occurred roughly in a dose-dependent manner. However, the level of endogenous hPrx 6 mRNA did not change in any group treated with the four extracts. The SOD activity was in agreement with the luciferase activity. Therefore, these results indicate that the hPrx 6/Luc vector system may successfully express and respond to antioxidant compounds in NCI-H460 cells. The data also suggest that the Prx 6/Luc vector system may be effectively applied in screening the response of hPrx 6 to antioxidant compounds in transgenic mice.

• Journal of Ginseng Research
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• v.21 no.3
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• pp.141-146
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• 1997

The role of cAMP in the differentiation process of F9 cells induced by ginsenosides was examined by performing transient transfixion assay with CRE-luciferase reporter plasmid, GR thansactivation assay with GRE-luciferase activity with or without treatment of CAMP and forskolin, an activator of adenylate cyclase, and protein klnase A assay in the presence of ginsenosides. Ginsenosides had no effect on CRE-transactivation activity, whereas retinoic acid induced the activity. When cAMP or forskolin was treated with ginsenosides, GRE-luciferase activity was further augumented by them. In addition, ginsenosides induced protein kinase A activity in the presence of cAMP. These results suggest that ginsenosides activate cAMP-dependent protein kinase A which, in turn, increase GR activity in F9 cells.

• BMB Reports
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• v.44 no.2
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• pp.102-106
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• 2011

Thermodynamic stability and refolding kinetics of firefly luciferase and three representative mutants with depletion of negative charge on a flexible loop via substitution of Glu by Arg (ER mutant) or Lys (EK mutant) as well as insertion of another Arg in ER mutants (ERR mutant) was investigated. According to thermodynamic studies, structural stability of ERR and ER mutants are enhanced compared to WT protein, whereas, these mutants become prone to aggregation at higher temperatures. Accordingly, it was concluded that enhanced structural stability of mutants depends on more compactness of folded state, whereas aggregation at higher temperatures in mutants is due to weakening of intermolecular repulsive electrostatic interactions and increase of intermolecular hydrophobic interactions. Kinetic results indicate that early events of protein folding are accelerated in mutants.

• BMB Reports
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• v.33 no.4
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• pp.353-357
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• 2000

A new assay method of ${\alpha}-Oxidase$ (fatty acid : oxygen dioxygenase, 1-decarboxylating) was developed using a bioluminescence reaction system of marine luminous bacterium, Photobacterium phosphoreum. ${\alpha}$-Oxidase was isolated from a cucumber (Cucumis sativus). Pentadecanoic acid was used as a substrate, and the product, tetradecanal, was analyzed with a bacterial luciferase-coupled reaction. Initial light intensity was directly related to the concentration of tetradecanal in the range of 1 nM to 10 ${\mu}M$. Optimal pH and temperature were 7.5 and $25^{\circ}C$, respectively. Optimal pentadecanoic acid concentration in a standard assay of ${\alpha}$-oxidase was 0.1 mM. The Km value of pentedecanoic acid was $85{\mu}M$. This method is straightforward, rapid, convenient, and easy. Its needs no treatment or extraction of reaction mixture.

• The Journal of Natural Sciences
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• v.15 no.1
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• pp.89-95
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• 2005

JSRV, which causes sheep lung cancer, is known to have the transforming activity of NIH3T3 cells. Especially Envelope protein of this virus has the transforming activity to NIH3T3. To know the effects on the transcriptional activity of transcription factors by this viral protein transient transfection was performed by using the luciferase reporter system. The result showed that JSRV Envelope protein increased the transcriptional activity of NF-kB and AP-1.