• Genomics & Informatics
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• v.8 no.1
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• pp.50-57
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• 2010

Mercuric chloride, a model nephrotoxicant was used to elucidate time- and dose- dependent global gene expression changes associated with proximal tubular toxicity. Rat kidney cell lines NRK-52E cells were exposed for 2, 6 and 12 hours and with 3 different doses of mercuric chloride. Cell viability assay showed that mercuric chloride had toxic effects on NRK-52E cells causing 20% cell death (IC20) at $40{\mu}M$ concentration. We set this IC20 as high dose concentration and 1/5 and 1/25 concentration of LC20 were used as mid and low concentration, respectively. Analyses of microarray data revealed that 738 genes were differentially expressed (more than two-fold change and p<0.05) by low concentration of mercuric chloride at least one time point in NRK-52E cells. 317 and 2,499 genes were differentially expressed at mid and high concentration of mercuric chloride, respectively. These deregulated genes showed a primary involvement with protein trafficking (CAV2, CANX, CORO1B), detoxification (GSTs) and immunity and defense (HMOX1, NQO1). Several of these genes were previously reported to be up-regulated in proximal tubule cells treated with nephrotoxicants and might be aid in promoting the predictive biomarkers for nephrotoxicity.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2001.06a
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• pp.77-80
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• 2001

A variety of different methods to generate diverse proteins, including random mutagenesis and recombination, are currently available, and most of them accumulate the mutations on the target gene of a protein, whose sequence space remains unchanged. On the other hand, a pool of diverse genes, which is generated by random insertions, deletions, and exchange of the homologous domains with different lengths in the target gene, would present the protein lineages resulting in new fitness landscapes. Here we report a method to generate a pool of protein variants with different sequence spaces by employing green fluorescent protein (GFP) as a model protein. This process, designated functional salvage screen (FSS), comprises the following procedures： a defective GFP template expressing no fluorescence is firstly constructed by genetically disrupting a predetermined region(s) of the protein, and a library of GFP variants is generated from the defective template by incorporating the randomly fragmented genomic DNA from E. coli into the defined region(s) of the target gene, followed by screening of the functionally salvaged, fluorescence-emitting GFPs. Two approaches, sequence-directed and PCR-coupled methods, were attempted to generate the library of GFP variants with new sequences derived from the genomic segments of E. coli. The functionally salvaged GFPs were selected and analyzed in terms of the sequence space and functional property. The results demonstrate that the functional salvage process not only can be a simple and effective method to create protein lineages with new sequence spaces, but also can be useful in elucidating the involvement of a specific region(s) or domain(s) in the structure and function of protein.

• The Plant Pathology Journal
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• v.29 no.1
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• pp.17-30
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• 2013

Barley stripe mosaic virus (BSMV) induces massive actin filament thickening at the infection front of infected Nicotiana benthamiana leaves. To determine the mechanisms leading to actin remodeling, fluorescent protein fusions of the BSMV triple gene block (TGB) proteins were coexpressed in cells with the actin marker DsRed: Talin. TGB ectopic expression experiments revealed that TGB3 is a major elicitor of filament thickening, that TGB2 resulted in formation of intermediate DsRed:Talin filaments, and that TGB1 alone had no obvious effects on actin filament structure. Latrunculin B (LatB) treat-ments retarded BSMV cell-to-cell movement, disrupted actin filament organization, and dramatically decreased the proportion of paired TGB3 foci appearing at the cell wall (CW). BSMV infection of transgenic plants tagged with GFP-KDEL exhibited membrane proliferation and vesicle formation that were especially evident around the nucleus. Similar membrane proliferation occurred in plants expressing TGB2 and/or TGB3, and DsRed: Talin fluorescence in these plants colocalized with the ER vesicles. TGB3 also associated with the Golgi apparatus and overlapped with cortical vesicles appearing at the cell periphery. Brefeldin A treatments disrupted Golgi and also altered vesicles at the CW, but failed to interfere with TGB CW localization. Our results indicate that actin cytoskeleton interactions are important in BSMV cell-to-cell movement and for CW localization of TGB3.

• Animal cells and systems
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• v.1 no.1
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• pp.151-155
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• 1997

Promoter of the Drosophila proliferating cell nuclear antigen (PCNA) gene contains DRE (Drosophila DNA replication-related element) required for the high level expression of replication-related genes. Recently, we found that promoter region of the D-raf (a Drosophila homolog of the human c-raf-1) contains two sequences homologous to the DRE and demonstrated the DRE/DREF (DRE-binding factor) involvement in regulation of the D-raf gene. In this study, using ursolic acid (UA), a pentacyclic triterpene acid reported to possess antitumor activities, we examined effects of UA on proliferation of the Drosophila cultured Kc cells and on expression of the PCNA and D-raf genes. UA showed an inhibitory effect on proliferation of the Kc cells in a concentration-dependent manner in DNA content assays and [3H]thymidine incorporation assays. The IC50 value of anti-proliferative effects of UA in DNA content assays was about 7.5uM. UA showed inhibitory effects on expression of the PCNA as well as on that of the D-raf, which were examined with the reporter plasmic p5'-168DPCNACAT or p5'-878DrafCAT, respectively. The results obtained in the present study suggest that expression of the PCNA and D-raf genes is coordinately regulated in at least UA-treated Kc cells and that down-regulation of expression of the PCNA and D-raf genes might be related with the antitumor activities of UA.

• Genomics & Informatics
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• v.7 no.1
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• pp.13-19
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• 2009

Osteoporosis is characterized by impaired osteogenesis. BMD is a major determinant of bone strength. The role of the VDR gene in predisposition to primary osteoporosis has been recognized. However, population-based case-control studies have been reported controversial results for known candidate genes in an ethnically distinct group. To determine the genetic effects of VDR variants on osteoporosis and BMD, we directly sequenced the VDR gene in 24 unrelated Korean individuals and identified eighteen sequence variants. We investigated the potential involvement of eight SNPs in osteoporosis in postmenopausal women (n = 729). Two SNPs (LD) in intron 2, -5294G>C (rs2238135) and -4817G>A (rs17882443) showed the evidence of association with enhanced BMD of the femoral neck ($p_{additive}$=0.031 for rs2238135; $p_{additive}$=0.017 and $p_{dominant}$= 0.019 for 17882443). Moreover, VDR -4817G>A was significantly associated with protective effect on all fracture risk ($p_{recessive}$=0.035, OR=0.2, 95% CI=$0.05{\sim}0.89$), and tended to be higher BMD values at various proximal femur sites. Therefore, we suggest that the -4817G>A may be useful genetic marker for vitamin D-related metabolism and may have an important role in the increased BMD of the proximal femur in postmenopausal Korean women.

• The Korean Journal of Physiology and Pharmacology
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• v.23 no.6
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• pp.529-537
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• 2019

Lung cancer is the most common cause of cancer deaths worldwide and several molecular signatures have been developed to predict survival in lung cancer. Increasing evidence suggests that proliferation and migration to promote tumor growth are associated with dysregulated ion channel expression. In this study, by analyzing high-throughput gene expression data, we identify the differentially expressed $K^+$ channel genes in lung cancer. In total, we prioritize ten dysregulated $K^+$ channel genes (5 up-regulated and 5 down-regulated genes, which were designated as K-10) in lung tumor tissue compared with normal tissue. A risk scoring system combined with the K-10 signature accurately predicts clinical outcome in lung cancer, which is independent of standard clinical and pathological prognostic factors including patient age, lymph node involvement, tumor size, and tumor grade. We further indicate that the K-10 potentially predicts clinical outcome in breast and colon cancers. Molecular signature discovered through $K^+$ gene expression profiling may serve as a novel biomarker to assess the risk in lung cancer.

• Plant Breeding and Biotechnology
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• v.5 no.3
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• pp.237-242
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• 2017

AT-hook proteins of plant have shown to be involved in growth and development through the modification of chromatin architecture to co-regulate transcription of genes. Recently, many genes encoding AT-hook protein have been identified and their involvement in senescence delay is investigated. In this study, soybean transgenic plants overexpressing chromatin architecture-controlling ATPG7 gene was produced by Agrobacterium-mediated transformation and investigated for the positive effect on the important agronomic traits mainly focusing on yield-related components. A total of 27 transgenic soybean plants were produced from about 400 explants. $T_1$ seeds were harvested from all transgenic plants. In the analysis of genomic DNAs from soybean transformants, ATPG7 and Bar fragments were amplified as expected, 975 bp and 408 bp in size, respectively. And also exact gene expression was confirmed by reverse transcriptase-PCR (RT-PCR) from transgenic line #6, #7 and #8. In a field evaluation of yield components of ATPG7 transgenic plants ($T_3$), higher plant height, more of pod number and greater average total seed weight were observed with statistical significance. The results of this study indicate that the introduction of ATPG7 gene in soybean may have the positive effect on yield components.

• Korean Journal of Breeding Science
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• v.42 no.5
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• pp.481-487
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• 2010

The ripening behavior of three apple cultivars, 'Tsugaru', 'Hongro' and 'Fuji' was distinctive and the involvement of POLYGALACTURONASE(PG) in the fruit softening process was confirmed to be ethylene dependent. Fruit softening is genetically coordinated by the action of several cell wall enzymes, including PG which depolymerizes cell wall pectin. Also, loss of firmness is associated with increasing of the ripening hormone, ethylene. In this work, climacteric ripening of three apple cultivars, Tsugaru, Hongro and Fuji, producing different ethylene levels and ripening responses, was examined. Correspondingly in Fuji, a linear and basal ethylene level was observed over the entire period of measurements, and Tsugaru and Hongro displayed a typical climacteric rise in ethylene production. Transcript accumulation of genes involved in ethylene biosynthesis (MdACS3 and MdACO1) and MdPG1 was studied in Tsugaru, Hongro and Fuji cultivars. Expression of MdACO1 transcripts was shown in all three ripened apple fruits. However, the MdACS3 and MdPG1 were transcribed differently in these cultivars. Comparing the MdPG1 of 'Tsugaru', 'Hongro' and 'Fuji', structural difference was discovered by genomic Southern analysis. Overall results pointed out that MdACS3 and MdPG1 play an important role in regulation of fruit ripening in apple cultivar.

• International Journal of Industrial Entomology and Biomaterials
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• v.43 no.1
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• pp.29-36
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• 2021

Silkworms have long been bred with human history to produce silk. It has been with humans for longer than other industrial insects, and the silkworm variety has been continuously improved. Silkworms have been developed into the optimal form for producing high quality silk and pupae. Recently, the production of transgenic silkworms has further expanded the possibility of industrial value of silkworms. Kynurenine 3-monooxygenase (KMO), which is a flavin enzyme, is known for its involvement in ommochrome pigment synthesis. In the field of mammals, including humans, previous studies have revealed the function and role of KMO, which is an important enzyme for various immune responses and cell protection. However, in the case of insects, the function of KMO has only been studied to be involved in the formation of pigment, and accordingly, KMO is used exclusively on screening for generation of transgenic insects as a marker. In this study, using KMO-edited silkworms, it was intended to discover the novel functions and roles of KMO in silkworms by identifying changes in the expression of various genes associated with stress and growth. The changes were observed in expressions of genes regulating on stresses to survive and those on ecdysteroid hormone between wild-type (WT) silkworms and kmo mutant silkworms. The loss of KMO, in particular, decreased the expression of the shadow (sad) gene, one of the Halloween genes in the synthesis of ecdysteroid. In conclusion, these results suggest that silkworm KMO is responsible for potential functions regarding stress response and ecdysteroid synthesis.

• Biomolecules & Therapeutics
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• v.30 no.3
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• pp.238-245
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• 2022

Previous reports have demonstrated that genetic mechanisms greatly mediate responses to drugs of abuse, including methamphetamine (METH). The circadian gene Period 2 (Per2) has been previously associated with differential responses towards METH in mice. While the behavioral consequences of eliminating Per2 have been illustrated previously, Per2 overexpression has not yet been comprehensively described; although, Per2-overexpressing (Per2 OE) mice previously showed reduced sensitivity towards METH-induced addiction-like behaviors. To further elucidate this distinct behavior of Per2 OE mice to METH, we identified possible candidate biomarkers by determining striatal differentially expressed genes (DEGs) in both drug-naïve and METH-treated Per2 OE mice relative to wild-type (WT), through RNA sequencing. Of the several DEGs in drug naïve Per2 OE mice, we identified six genes that were altered after repeated METH treatment in WT mice, but not in Per2 OE mice. These results, validated by quantitative real-time polymerase chain reaction, could suggest that the identified DEGs might underlie the previously reported weaker METH-induced responses of Per2 OE mice compared to WT. Gene network analysis also revealed that Asic3, Hba-a1, and Rnf17 are possibly associated with Per2 through physical interactions and predicted correlations, and might potentially participate in addiction. Inhibiting the functional protein of Asic3 prior to METH administration resulted in the partial reduction of METH-induced conditioned place preference in WT mice, supporting a possible involvement of Asic3 in METH-induced reward. Although encouraging further investigations, our findings suggest that these DEGs, including Asic3, may play significant roles in the lower sensitivity of Per2 OE mice to METH.