• Journal of Life Science
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• v.21 no.9
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• pp.1301-1309
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• 2011

The major function of adipocytes is to store fat in the form of triglycerides. One of the signaling pathways known to affect adipogenesis, i.e. fat formation, is the WNT/${\beta}$-catenin pathway which inhibits the expression and activity of key regulators of adipogenesis. The purpose of this research is to find genes among the WNT/${\beta}$-catenin pathway which regulate adipogenesis by using small interfering (si) RNA and to find the association of single nucleotide polymorphisms (SNPs) of the gene with serum triglyceride levels in the human population. To elucidate the effects of ${\beta}$-catenin siRNA on adipogenesis key factors, PPAR${\gamma}$ and C/EBP${\alpha}$, we performed real-time PCR and western blotting experiments for the analyses of mRNA and protein levels. It was found that the siRNA-mediated knockdown of ${\beta}$-catenin upregulates adipogenesis key factors. However, upstream regulators of the WNT/${\beta}$-catenin pathway, such as DVL2 and LRP6, had no significant effects compared to ${\beta}$-catenin. These results indicate that ${\beta}$-catenin is a candidate gene for human fat accumulation. In general, serum triglyceride level is a good indicator of fat accumulation in humans. According to statistical analyses of the association between serum triglyceride level and SNPs of ${\beta}$-catenin, -10,288 C>T SNP (rs7630377) in the promoter region was significantly associated with serum triglyceride levels (p<0.05) in 290 Korean subjects. On the other hand, serum cholesterol levels were not significantly associated with SNPs of the ${\beta}$-catenin gene. The results of this study showed that ${\beta}$-catenin is associated with fat accumulation both in vitro and in the human population.

• Horticultural Science & Technology
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• v.28 no.3
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• pp.449-455
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• 2010

'Moulinrouge' was selected as the best regenerating cultivar among 18 different spray-type chrysanthemum cultivars bred in the Gyeongnam Flowers Breeding Research Institute. When the leaf explants from standard- and spray-type chrysanthemum 'Jinba' and 'Moulinrouge' were incubated on MS basal medium supplemented with $0.5mg{\cdot}L^{-1}$ BA and $1.0mg{\cdot}L^{-1}$ NAA, both 'Jinba' and 'Moulinrouge' induced adventitious shoots that can be regenerated into plantlets. Based on these regeneration conditions, we developed an efficient $Agrobacterium$-mediated chrysanthemum 'Moulinrouge' transformation method by using sequential selection of shoots from low ($10mg{\cdot}L^{-1}$) to high ($30mg{\cdot}L^{-1}$) concentrations of kanamycin after co-cultivation of leaf explants with $Agrobacterium$ for 10 days and induction of shoots. All kanamycin resistant plants investigated with genomic PCR analysis carried the report gene, $AtSICKLE$, in their genome. Although expression levels of the report gene in the transgenic plants investigated with RT-PCR were relatively low because of inefficiency of CaMV 35S promoter in chrysanthemum, transgenic lines expressing $AtSICKLE$ efficiently showed leaf epinasty phenotype. We expect that our results will provide a useful method that can perform a high-throughput investigation of genes isolated and studied well in model plants for molecular breeding of chrysanthemum.

• Tuberculosis and Respiratory Diseases
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• v.57 no.3
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• pp.265-272
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• 2004

Background : RASSF1A, which is one of tumor suppressor genes, is frequently inactivated by hypermethylation of the promoter region in a variety of human cancers, including lung cancer. This study was performed to investigate the association between RASSF1A methylation and the clinicopathological factors in patients with squamous cell carcinoma of the lung. Methods : Eighty-one samples from the patients with squamous cell carcinoma of lung were examined. The promoter methyation of RASSF1A was analyzed by methylation specific PCR and sequencing. Statistical analysis was made to examine the association between RASSF1A methylation and the clinicopathological parameters. Results : RASSF1A methylation was observed in 37.0 % (30 of 81) of the patients with squamous cell carcinoma of the lung. RASSF1A methylation was found to be associated with cellular differentiation(p=0.0097) and the overall survival(p=0.0635). However, there was no association between RASSF1A methylation and the other clinicopathological parameters, such as the pathological TNM stage, the recurrence rate, lymph node invasion and the amount of cigarettes smoked. Conclusion : RASSF1A methylation might be associated with a poor prognosis in patients with squamous carcinoma of the lung. A larger scale study is needed.

• Journal of Life Science
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• v.16 no.6
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• pp.1052-1059
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• 2006

The heat shock response is induced by environmental stress, pathophysiological state and non-stress conditions and wide spread from bacteria to human. Although translations of most proteins are stopped under a heat shock response, heat shock proteins (HSPs) are produced to protect cell from stress. When heat shock response is induced, conformation of HSF1 was changed from monomer to trimer and HSF1 specifically binds to DNA, which was called a heat shock element(HSE) within the promoter of the heat shock genes. Human HSF1(hHSFl) contains five cysteine(Cys) residues. A thiol group(R-SH) of Cys is a strong nucleophile, the most readily oxidized and nitrosylated in amino acid chain. This consideration suggests that Cys residues may regulate the change of conformation and the activity of hHSF1 through a redox-dependent thiol/disulfide exchange reaction. We want to construct role of five Cys residues of hHSF by redox reagents. According to two studies, Cys residues are related to trimer formation of hHSF1. In this study, we want to demonstrate the correlation between structural change and DNA-binding activity of HSF1 through forming disulfide bond and trimerization. In this results, we could deduce that DNA binding activity of DNA binding domain wasn't affected by redox for always expose outside to easily bind to DNA. DNA binding activity of wild-type HSF's DNA binding domain was affected by conformational change, as conformational structure change (trimerization) caused DNA binding domain.

• Journal of Life Science
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• v.19 no.7
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• pp.949-955
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• 2009

Mucosal epithelia sense external stress signals and transmit them to the intracellular cascade responses. Ribotoxic stress-producing chemicals such as deoxynivalenol (DON) or other trichothecene mycotoxins have been linked with gastrointestinal inflammatory diseases by Fusarium-contamination. The purpose of this study was to test the hypothesis that DON evokes the epithelial sentinel signals of RNA-dependent protein kinase (PKR) and early growth response gene 1 (EGR-1), which together contribute to the pro-inflammatory cytokine interleukin 8 (IL-8) in human intestinal epithelial cells. PKR suppression by the dominant negative PKR expression attenuated DON-stimulated interleukin-8 production. Moreover, 1L-8 transcriptional activation by DON was also reduced by PKR inhibition in the human intestinal epithelial cells. Treatment with the PKR inhibitor also suppressed EGR-1 promoter activity, mRNA and protein induction, although mitogen-activated protein (MAP) kinases such as extracellular signal-regulated protein kinases (ERK) 1/2, p38, c-Jun N-terminal Kinase (INK) were little affected or even enhanced in presence of a PKR inhibitor. These patterns were also compared in the EGR-1-suppressed cells, which showed much more suppressed production of 1L-8. All things taken into consideration, DON-activated sentinel signals of EGR-1 via PKR mediated interleukin-8 production in human intestinal epithelial cells, which provide insight into the possible general mechanism associated with mucosal inflammation as an intestinal toxic insult by ribotoxic trichothecene mycotoxins.

• Journal of Plant Biotechnology
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• v.5 no.3
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• pp.143-148
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• 2003

Bax, a mammalian pro-apoptotic member of the Bcl-2 family induces cell death when expressed in yeast. To investigate whether Bax expression can induce cell death in plant, we produced transgenic Arabidopsis plants that contained murine Bax cDNA under control of a glucocorticoid-inducible promoter. Transgenic plants treated with dexamethasone, a strong synthetic glucocorticoid, induced Bax accumulation and cell death, suggesting that some elements of cell death mechanism by Bax may be conserved among various organisms. Therefore, we developed novel yeast genetic system, and cloned several Plant Bax Inhibitors (PBIs). Here, we report the function of two PBIs in detail. PBI1 is ascorbate peroxidase (sAPX). Fluorescence method of dihydrorhodamine123 oxidation revealed that expression of Bax in yeast cells generated reactive oxygen species (ROS), and which was greatly reduced by co-expression with sAPX. These results suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Baxinduced cell death in yeast. PBI2 encodes nucleoside diphosphate kinase (NDPK). ROS stress strongly induces the expression of the NDPK2 gene in Arabidopsis thaliana (AtNDPK2). Transgenic plants overexpressing AtNDPK2 have lower levels of ROS than wildtype plants. Mutants lacking AtNDPK2 had higher levels of ROS than wildtype. $H_2O_2$ treatment induced the phosphorylation of two endogenous proteins whose molecular weights suggested they are AtMPK3 and AtMPK6. In the absence of $H_2O_2$ treatment, phosphorylation of these proteins was slightly elevated in plants overexpressing AtNDPK2 but markedly decreased in the AtNDPK2 deletion mutant. Yeast two-hybrid and in vitro protein pull-down assays revealed that AtNDPK2 specifically interacts with AtMPK3 and AtMPK6. Furthermore, AtNDPK2 also enhances the MSP phosphorylation activity of AtMPK3 in vitro. Finally, constitutive overexpression of AtNDPK2 in Arabidopsis plants conferred an enhanced tolerance to multiple environmental stresses that elicit ROS accumulation in situ. Thus, AtNDPK2 appears to playa novel regulatory role in $H_2O_2$-mediated MAPK signaling in plants.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.8
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• pp.1131-1140
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• 2014

This research focused on the effects of different doses of Bacillus subtilis KN-42 on the growth performance, diarrhea incidence, faecal bacterial flora, and the relative number of Lactobacillus and Escherichia coli in faeces of weaned piglets to determine whether the strain can serve as a candidate antimicrobial growth promoter. A total of 360 piglets (initial body weight $7.14{\pm}0.63$ kg) weaned at $26{\pm}2$ days of age were randomly allotted to 5 treatment groups (4 pens per treatment with 18 pigs per pen) for a 28-day trial. Dietary treatments were basal diet without any antimicrobial (negative control; NC), basal diet supplemented with 120 mg/kg feed of neomycin sulfate (positive control; PC) and basal diet supplemented with $2{\times}10^9$ (L), $4{\times}10^9$ (M) and $20{\times}10^9$ (H) CFU/kg feed of B. subtilis KN-42. During the overall period, average daily gain and feed efficiency of piglets were higher in groups PC, M, and H than those in group NC (p<0.05), and all probiotics and antibiotics groups had a lower diarrhea index than group NC (p<0.05). The 16S rDNA gene-based methods were used to analyze faecal bacterial flora on day 28 of experiment. The result of denaturing gradient gel electrophoresis analysis showed that supplementation of B. subtilis KN-42 to the diet changed the bacterial communities, with a higher bacterial diversity and band number in group M than in the other four groups. Real-time polymerase chain reaction analysis showed that the relative number of Lactobacillus were higher in groups PC and H than in group NC (p<0.05), and the supplemented B. subtilis KN-42 to the diet also reduced the relative number of E. coli (p<0.05). These results suggest that dietary addition of B. subtilis KN-42 can improve the growth performance and gastrointestinal health of piglets.

• Korean Journal of Animal Reproduction
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• v.27 no.1
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• pp.9-14
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• 2003

The pBT-L transgenic mice carrying human TPO gene in conjunction with bovine $\beta$-casein promoter express human TPO in milk during lactation. In this study, stability of germ line transmission and expression of pBT-L transgene integrated into host chromosome were monitored up to generation F10 of transgenic pBT-L/15 line. When male mouse of generation F8 was crossbred with normal females, approximately half of offsprings (51.3$\pm$18.98%) were identified as transgenic mice. Generation F9 and F 10 mice also showed similar transmission rates (43.8$\pm$18.98% and 71.4$\pm$26.98%, respectively), implying that pBT-L transgene can be transmitted stably up to long term generation in the transgenic mice. Expression levels of human TPO from milk of generation F9 and F10 mice were 1.1$\pm$0.33 mg/ml and 1.1 $\pm$0.45 mg/ml, respectively, which are similar to expression level of generation F2 mice. In conclusion, our results suggest that transgenic animals once established will continuously pass their transgenes to the progeny through the breeding program with the same productivity of human protein in their milk.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2003.04a
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• pp.65-71
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• 2003

Bax, a mammalian pro-apoptotic member of the Bcl-2 family, induces cell death when expressed in yeast. To investigate whether Bax expression can induce cell death in plant, we produced transgenic Arabidopsis plants that contained murine Bax cDNA under control of a glucocorticoid-inducible promoter. Transgenic plants treated with dexamethasone, a strong synthetic glucocorticoid, induced Bax accumulation and cell death, suggesting that some elements of cell death mechanism by Bax may be conserved among various organisms. Therefore, we developed novel yeast genetic system, and cloned several Plant Bax Inhibitors (PBIs). Here, we report the function of two PBIs in detail. PBI1 is ascorbate peroxidase (sAPX). Fluorescence method of dihydrorho-damine 123 oxidation revealed that expression of Bax in yeast cells generated reactive oxygen species (ROS), and which was greatly reduced by co-expression with sAPX. These results suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Baxinduced cell death in yeast. PBI2 encodes nucleoside diphosphate kinase (NDPK). ROS stress strongly induces the expression of the NDPK2 gene in Arabidopsis thaliana (AtNDPK2). Transgenic plants overexpressing AtNDPK2 have lower levels of ROS than wildtype plants. Mutants lacking AtNDPK2 had higher levels of ROS than wildtype. $H_2O_2$ treatment induced the phosphorylation of two endogenous proteins whose molecular weights suggested they are AtMPK3 and AtMPK6. In the absence of $H_2O_2$ treatment, phosphorylation of these proteins was slightly elevated in plants overexpressing AtNDPK2 but markedly decreased in the AtNDPK2 deletion mutant. Yeast two-hybrid and in vitro protein pull-down assays revealed that AtNDPK2 specifically interacts with AtMPK3 and AtMPK6. Furthermore, AtNDPK2 also enhances the MBP phosphorylation activity of AtMPK3 in vitro. Finally, constitutive overexpression of AtNDPK2 in Arabidopsis plants conferred an enhanced tolerance to multiple environmental stresses that elicit ROS accumulation in situ. Thus, AtNDPK2 appears to play a novel regulatory role in $H_2O_2$-mediated MAPK signaling in plants.

• Journal of Microbiology and Biotechnology
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• v.19 no.10
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• pp.1223-1229
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• 2009

To construct a new recombinant strain of Bacillus velezensis that has antifungal and insecticidal activity via the expression of the insecticidal Bacillus thuringiensis crystal protein, a B. thuringiensis expression vector (pHT1K-1Ac) was generated that contained the B. thuringiensis cry1Ac gene under the control of its endogenous promoter in a minimal E. coli-B. thuringiensis shuttle vector (pHT1K). This vector was introduced into a B. velezensis isolate that showed high antifungal activities against several plant diseases, including rice blast (Magnaporthe grisea), rice sheath blight (Rhizotonia solani), tomato gray mold (Botrytis cinerea), tomato late blight (Phytophthora infestans), and wheat leaf rust (Puccinia recondita), by electroporation. The recombinant B. velezensis strain was confirmed by PCR using cry1Ac-specific primers. Additionally, the recombinant strain produced a protein approximately 130 kDa in size and parasporal inclusion bodies similar to B. thuringiensis. The in vivo antifungal activity assay demonstrated that the activity of the recombinant B. velezensis strain was maintained at the same level as that of wild-type B. velezensis. Furthermore, it exhibited high insecticidal activity against a lepidopteran pest, Plutella xylostella, although its activity was lower than that of a recombinant B. thuringiensis strain, whereas wild-type B. velezensis strain did not show any insecticidal activity. These results suggest that this recombinant B. velezensis strain can be used to control harmful insect pests and fungal diseases simultaneously in one crop.