• Proceedings of the Botanical Society of Korea Conference
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• 1996.07a
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• pp.61-74
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• 1996

Human and monogastric animals can not synthesize 10 out of the 20 amino asids and therefor need to obtain these from their diet. The plant seed is a major source of dietary protein. It is particular important in their study to increase nutritional quality of the seed storage proteins. The low contents of lysine, asparagine and threonenein various cereal seeds and of cystein and methionine. In legume seeds is due to the low proportions of these amino acids in the major storage proteins, we have tried to apply the three strategies; (1) mutagenesis and selection of specific amino acid analogue resistance, (2) cloning and expression study of lysine biosynthesis related gene, (3) transfomation of lysine rich soybean glycinin gene. The 5-methyltryptophan (5MT) resistant cell lines, SAR1, SAR2 and SAR3 were selected from anther derived callus of rice (Oryza sativa L. "Sasanishiki"). Among these selected cell lines, two (SAR1 and SAR3) were able to grow stably at 200 mg/L of 5MT. Analysis of the freed amino acids in callus shows that 5MT resistant cells (SAR3) accumulated free tryptophan at least up to 50 times higher than those that of the higher than of SAS. These results indicated that the 5MT resistant cell lines are useful in studies of amino acid biosynthesis. Tr75, a rice (Oryza sativa L., var. Sasanishiki) mutant resistant to 5MT was segregated from the progenies of its initial mutant line, TR1. The 5MT resistant of TR75 was inherited in the M8 generations as a single dominant nuclear gene. The content of free amino acids in the TR75 homozygous seeds increased approximately 1.5 to 2.0 fold compared to wild-type seeds. Especially, the contents of tryptophan, phenylalanine and aspartic acid were 5.0, 5.3 and 2.7 times higher than those of wild-type seeds, respectively. The content of lysine is significantly low in rice. The lysine is synthesized by a complex pathway that is predominantly regulated by feedback inhibition of several enzymes including asparginase, aspatate kinase, dihydrodipicolinat synthase, etc. For understanding the regulation mechanism of lysine synthesis in rice, we try to clone the lysine biosynthetic metabolism related gene, DHPS and asparaginase, from rice. We have isolated a rice DHPS genomic clone which contains an ORF of 1044 nucleotides (347 amino acids, Mr. 38, 381 daltons), an intron of 587 nucleotides and 5'and 3'-flanking regions by screening of rice genomic DNA library. Deduced amino acid sequence of mature peptide domain of GDHPS clone is highly conserved in monocot and dicot plants whereas that of transit peptide domain is extremely different depending on plant specie. Southern blot analysis indicated that GDHPS is located two copy gene in rice genome. The transcripts of a rice GDHPS were expressed in leaves and roots but not detected in callus tissues. The transcription level of GDHPS is much higher in leaves indicating enormous chloroplast development than roots. Genomic DNA clones for asparaginase genes were screened from the rice genomic library by using plaque hybridization technique. Twelve different genomic clones were isolated from first and second screening, and 8 of 12 clones were analyzed by restriction patterns and identified by Southern Blotting, Restriction enzyme digestion patterns and Southern blot analysis of 8 clones show the different pattern for asparaginase gene. Genomic Southern blot analysis from rice were done. It is estimated that rice has at least 2-3 copy of asparaginase gene. One of 8 positive clones was subcloned into the pBluescript SK(+) vector, and was constructed the physical map. For transformation of lysine rich storage protein into tobacco, soybean glycinin genes are transformed into tobacco. To examine whether glycinin could be stably accumulated in endosperm tissue, the glycinin cDNA was transcriptionally fused to an endosperm-specific promotor of the rice storage protein glutelin gene and then introduced into tobacco genomic via Agrobacterium-mediated transformation. Consequently the glycinin gene was expressed in a seed-and developmentally-specific manner in transgenic tobacco seeds. Glycinin were targeted to vacuole-derived protein bodies in the endosperm tissue and highly accumulated in the matrix region of many transgenic plant (1-4% of total seed proteins). Synthesized glycinin was processed into mature form, and assembled into a hexamer in a similar manner as the glycinin in soybean seed. Modified glycinin, in which 4 contiguous methionine residues were inserted at the variable regions corresponding to the C - teminal regions of the acidic and basic polypeptides, were also found to be accumulated similarly as in the normal glycinin. There was no apparent difference in the expression level, processing and targeting to protein bodies, or accumulation level between normal and modified glycinin. glycinin.

• Journal of Life Science
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• v.26 no.2
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• pp.147-154
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• 2016

Osteosarcoma (OS) is the most common and malignant bone tumors. Although many types of resection surgery and experimental agents were developed, median survival and clinical prognosis are poorly investigated. Recently, several researches have reported that Eucheuma cottonii has potent as protective effects of coal dust-induced lung damage via inhibition of malondialdehyde (MDA) and oxidative stress in bronchoalveolar lavage fluids (BALF). However, anti-cancer effects and specific molecular mechanism of extract from Eucheuma cottonii (EE) has not been clearly studied yet. This study evaluated that anti-cancer potential of EE in human osteosarcoma Saos-2 cells. EE indicated cytotoxicity on Saos-2 cells in a dose-dependent manner. Morphological degradation and nucleic condensation were also observed under the EE treatment. However, it did not significantly affect on non-cancerous kidney HEK-293 cells under the same concentration which is shown cytotoxicity on Saos-2 cells. The phosphorylation of Fas-Associated Death Domain (FADD) and expression of cleaved caspase-8, -7 and -3 were upregulated in a dose-dependent manner. In immunofluorescence staining, expression level of Fas and cleaved PARP were upregulated by EE treatment. Furthermore, treatment of EE induces upregulation of sub G1 phase by flow cytometry analysis. The results demonstrated that EE has a therapeutic potential against osteosarcoma via FADD mediated caspase cascade apoptosis signal pathway.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.45 no.2
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• pp.175-184
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• 2019

The epidermal growth factor (EGF) has a intrinsic function of inducing growth and proliferation of cells through interacting with cell membrane receptors in human epidermis and dermis layer. These functions of EGF are used as a main ingredient for wound healing medicines and anti-aging cosmetics. As a cosmetic ingredient, the EGF has a problem in exhibiting its natural efficacy due to the lack of the ability to penetrate through the stratum corneum, which is known as the skin barrier. In this study, a recombinant human epidermal growth factor ($MTD_{151}-EGF$) fused with the macromolecule transduction domain $(MTD)_{151}$ with the skin penetration ability was developed to improve the skin penetration efficiency of the EGF. Expression of $MTD_{151}-EGF$ was performed in E. coli transformed with a vector encoding the $MTD_{151}-EGF$ gene and then purified. The purified $MTD_{151}-EGF$ was evaluated using cell proliferation assay, cytotoxicity test and skin penetration test by franz diffusion cell assay and artificial skin. Cell proliferation activity of $MTD_{151}-EGF$ purified to high purity of 99% or above was equivalent to the EGF or better, and cytotoxicity was not observed. In addition, the $MTD_{151}-EGF$ showed an excellent penetration efficiency compared to the EGF in the skin penetration test with EGF and $MTD_{151}-EGF$ labeled by FITC in an artificial skin penetration model. Based on the quantitative analysis of the penetrating substance using franz diffusion cell assay, the amount of penetration was about 16 times more than that of EGF. These results can be regarded as an effective alternative to improve the existing physical transdermal penetration method related to the use of various active ingredients for cosmetics.

• Clinical and Experimental Pediatrics
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• v.45 no.2
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• pp.183-191
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• 2002

Purpose : X-linked agammaglobulinemia(XLA) is an immunodeficiency caused by abnormalities in Bruton's tyrosine kinase(Btk), and is characterized by a deficiency of peripheral blood B cells. We studied the cytoplasmic expression of Btk protein and analyzed the Btk gene in peripheral blood mononuclear cells from two siblings and one cousin with XLA, as well as additional family members. Methods : Btk protein expression was analyzed by flow cytometry. Isolation of the coding sequence of the Btk gene was performed by amplification using the reverse transcription-polymerase chain reaction(RT-PCR) technique. Sequence alterations were screened by the single-stranded conformation polymorphism(SSCP) method and characterized by standard sequencing protocols. Results : Cytoplasmic expression of Btk protein in monocytes was not detected in three patients with XLA. In addition, Btk protein analysis clearly showed cellular mosaicism in monocytes from four obligate carriers, findings further supported by SSCP. A single base pair mutation(T to C) in Btk-exon three, which encodes the PH domain, was identified in four XLA patients. A diagnostic sequencing analysis was established to detect heterozygotic pattern in 4 carrier females. Furthermore, we found significant clinical heterogeneity in individuals with the same gene mutation. Conclusion : The implicating genetic alteration provided valuable clues to the pathogenesis of XLA in Korea and the flow cytometric analysis was suggested as a useful tool for rapid detection of XLA patients and carriers. The present study has identified a genetic mutation in the Btk coding region and demonstrated heterogeneity in clinical manifestations among patients with the same mutation. A flow cytometric analysis was found to be informative in establishing a deficiency of Btk protein in both patients and carriers and is recommended as a frontline procedure in the molecular diagnosis and work-up of XLA.

• Cartoon and Animation Studies
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• s.48
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• pp.359-383
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• 2017

3,000 years ago, since the first poster of humanity appeared in Egypt, the invention of printing technique and the development of civilization have accelerated the poster production technology. In keeping with this, the expression of poster has also been developed as an attempt to express artistic sensibility in a simple arrangement of characters, and now it has become an art form that has become a domain of professional designers. However, the technological development in the expression of poster is keep staying in two-dimensional, and is dependent on printing only that it is irrelevant to the change of ICT environment based on modern multimedia. Especially, among the many kinds of posters, the style of movie posters, which are the only objects for video, are still printed on paper, and many attempts have been made so far, but the movie industry still does not consider ICT integration at all. This study started with the feature that the object of the movie poster dealt with the video and attempted to introduce the augmented reality to apply the dynamic image of the movie to the static poster. In the graduation work of the media design major of a university in Korea, the poster of each works for promoting the visual work of the students was designed and printed in the form of a commercial film poster. Among them, 6 artworks that are considered to be suitable for augmented reality were selected and augmented reality was introduced and exhibited. Content that appears matched to the poster through the mobile device is reproduced on a poster of a scene of the video, but the text informations of the original poster are kept as they are, so that is able to build a moving poster looked like a wanted from the movie "Harry Potter". In order to produce this augmented reality poster, we applied augmented reality to posters of existing commercial films produced in two different formats, and found a way to increase the characteristics of AR contents. Through this, we were able to understand poster design suitable for AR representation, and technical expression for stable operation of augmented reality can be summarized in the matching process of augmented reality contents production.

• Journal of Life Science
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• v.19 no.1
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• pp.101-110
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• 2009

The regulation of gene expression plays an important role in cell cycle controls. In this study, a novel $mas2^+$ (mitosis associated protein) gene, a homolog of human SMARCAD1 was isolated and characterized from a fission yeast Schizosaccharomyces pombe (S. pombe) using gene-specific polymerase chain reaction. The isolated gene contained a complete open reading frame capable of encoding 922 amino acid residues with a typical promoter, as judged by nucleotide sequence analysis. It was also found that an SNF2 domain is located, which is involved in the chromosome remodeling. The quantitative analysis of the $mas2^+$ transcript against $adh1^+$ showed that the expression level of $mas2^+$ is high before septum formation in S. pombe. When $mas2^+$ null mutant cells were grown at 27 and $35^{\circ}C$, the cytokinesis of $mas2^+$ null mutant was greatly delayed and a large number of multi-septate and mis-segregated cells were produced. In addition, the number of multi-septate cells significantly increased. When cells were cultured in YES rich medium to increase proliferation, the abnormal phenotypes $mas2^+$ null mutant dramatically increased. These phenotypes could be rescued by an over-expression of the mast gene. The Mas2 protein localized in the nuclei of S. pombe, as evidenced by Mas2-EGFP signals. These results suggest that the $mas2^+$ is homologous to human SMARCAD1 gene and involved in septum formation and chromosome remodeling control.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.1
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• pp.33-46
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• 2005

연구목적: ADAMs은 metalloprotease/disintegrin domain을 가진 transmemebrane glycoprotein으로써 지금까지 30개 이상의 ADAM 및 10개 이상의 ADAMTS가 알려져 있다. 이들의 기능은 포유동물의 수정 시 sperm-egg binding과 fusion, myoblast fusion, integrin과의 결합 등에 직접 관여하거나, TNF-alpha 등의 생체신호전달물질이 세포로부터 분비될 때에 이들의 구조를 변화시켜 활성화시키는 효소작용, 그리고 dendritic cell differentiation 등에 관여하는 것으로 알려져 있다. 그러나 자궁내막 조직에서의 유전자 및 단백질 발현 여부에 관해서는 거의 보고되어 있지 않고 있다. 본 연구에서는 착상 전후 시기의 생쥐 자궁조직에서 ADAM-8, 9, 10, 12, 15, 17 그리고 ADAMTS-1의 유전자가 발현하는 지를 알아보았다. 연구 재료 및 방법: 본 연구에서는 생쥐의 자궁조직을 대상으로 ADAM-8, 9, 10, 12, 15, 17 그리고 ADAMTS-1을 선정하여, 초기 임신 기간에서의 유전자 발현 여부를 조사하였고 이 결과를 바탕으로 자궁조직에서의 이들 유전자들의 생리적인 기능을 규명하고자 하였다. 결 과: 임신한 생쥐 자궁조직에서의 ADAM-8, 9, 10, 12, 15, 17 그리고 ADAMTS-1의 유전자 및 단백질의 발현 양상을 RT-PCR 방법을 이용하여 알아본 결과, 조사된 ADAM 종류와 임신 날짜별로 다르게 나타났다. ADAM-8의 유전자 전사체는 임신 1일째 매우 강하게 발현되었으나 임신 3일째로 진행되면서 감소하다가 이후 다시 임신 5일째가 되면서 증가하는 양상을 보였다. ADAM-9, 10, 17 그리고 ADAMTS-1의 경우는 임신 1일째에서 5일째까지 유전자의 발현 양상이 크게 변하지 않았고 ADAM-12와 ADAM-15의 유전자 전사체는 임신 1일에서 5일로 진행되면서 현저하게 증가되는 양상을 보였다. 이후 임신 6일에서 8일에서는 생쥐 배아가 착상된 부위와 비 착상부위로 나누어 유전자의 발현 양상을 관찰한 결과, 조사된 ADAM 모두 비착상 부위보다 착상부위에서 유전자 전사체의 발현이 크게 증가되는 것으로 나타났다. 결 론: 이상의 결과로 미루어 ADAM 유전자는 임신초기 착상과정과 임신 단계에 따른 자궁의 조직 재구성에 중요한 역할을 할 것으로 생각된다.

• Journal of Life Science
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• v.30 no.9
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• pp.826-833
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• 2020

Phospholipase C gamma (PLCγ) has critical roles in receptor tyrosine kinase- and non-receptor tyrosine kinase-mediated cellular signaling relating to the hydrolysis of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P₂] to produce inositol 1,4,5 trisphosphate (IP₃) and diacylglycerol (DAG), which promote protein kinase C (PKC) and Ca²⁺ signaling to their downstream cellular targets. PLCγ has two isozymes called PLCγ1 and PLCγ2, which control cell growth and differentiation. In addition to catalytically active X- and Y-domains, both isotypes contain two Src homology 2 (SH2) domains and an SH3 domain for protein-protein interaction when the cells are activated by ligand stimulation. PLCγ also contains two pleckstrin homology (PH) domains for membrane-associated phosphoinositide binding and protein-protein interactions. While PLCγ1 is widely expressed and appears to regulate intracellular signaling in many tissues, PLCγ2 expression is restricted to cells of hematopoietic systems and seems to play a role in the regulation of immune response. A distinct mechanism for PLCγ activation is linked to an increase in phosphorylation of specific tyrosine residue, Y783. Recent studies have demonstrated that PLCγ mutations are closely related to cancer, immune disease, and brain disorders. Our review focused on the physiological roles of PLCγ by means of its structure and enzyme activity and the pathological functions of PLCγ via mutational analysis obtained from various human diseases and PLCγ knockout mice.

• Journal of Life Science
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• v.28 no.7
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• pp.765-771
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• 2018

Peroxisome proliferator-activated receptor gamma ($PPAR{\gamma}$) is a key transcription factor that regulates adipogenesis, and epigenetic control of $PPAR{\gamma}$ is of great interest in obesity-inhibition research. Our previous study showed that CACUL1 (CDK2-associated cullin domain 1) acts as a corepressor that inhibits $PPAR{\gamma}$ transcriptional activity and adipocyte differentiation. Here, we investigated the roles of protein arginine methyltransferase 5 (PRMT5), a novel binding partner of CACUL1, in regulating $PPAR{\gamma}$. The interaction between PRMT5 and CACUL1 was shown by immunoprecipitation assay in vivo and GST pulldown assay in vitro. As shown by luciferase reporter assay, PRMT5 and CACUL1 cooperated to inhibit the transcriptional activity of $PPAR{\gamma}$. The suppressive role of PRMT5 in adipogenesis was examined by Oil Red O staining using 3T3-L1 cells, which stably overexpress or deplete PRMT5. Overexpression of PRMT5 suppresses $PPAR{\gamma}$-mediated adipogenesis, whereas PRMT5 knockdown increases lipid accumulation in 3T3-L1 cells. Consistently, PRMT5 attenuates the expression of Lpl and aP2, the target genes of $PPAR{\gamma}$, as demonstrated by RT-qPCR analysis. Overall, these results suggest that PRMT5 interacts with CACUL1 to impair the transcriptional activity of $PPAR{\gamma}$, leading to the inhibition of adipocyte differentiation. Therefore, the regulation of PRMT5 enzymatic activity may provide a clue to develop an anti-obesity drug.

• Journal of Life Science
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• v.25 no.10
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• pp.1103-1109
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• 2015

Cholinergic innervation of the hippocampus is known to be correlated with learning and memory. The cholinergic agonist carbachol (CCh) modulate synaptic plasticity and produced long-term synaptic depression (LTD) in the hippocampus. However, the exact mechanisms by which the cholinergic system modifies synaptic functions in the hippocampus have yet to be determined. This study introduces an acetylcholine receptor-mediated LTD that requires internalization of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors on the postsynaptic surface and their intracellular mechanism in the hippocampus. In the present study, we showed that the application of the cholinergic agonist CCh reduced the surface expression of GluA2 on synapses and that this reduction was prevented by the M1 muscarinic acetylcholine receptor antagonist pirenzepine in primary hippocampal neurons. The interaction between GluA2 and the glutamate receptor-interacting protein 1 (GRIP1) was disrupted in a hippocampal slice from a rat upon CCh simulation. Under the same conditions, the binding of GluA2 to adaptin-α, a protein involved in clathrin-mediated endocytosis, was enhanced. The current data suggest that the activation of LTD, mediated by the acetylcholine receptor, requires the internalization of the GluA2 subunits of AMPA receptors and that this may be controlled by the disruption of GRIP1 in the PDZ ligand domain of GluA2. Therefore, we can hypothesize that one mechanism underlying the LTD mediated by the M1 mAChR is the internalization of the GluA2 AMPAR subunits from the plasma membrane in the hippocampal cholinergic system.