• Journal of Ginseng Research
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• v.41 no.3
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• pp.403-410
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• 2017

Background: Prenyltransferases catalyze the sequential addition of isopentenyl diphosphate units to allylic prenyl diphosphate acceptors and are classified as either trans-prenyltransferases (TPTs) or cis-prenyltransferases (CPTs). The functions of CPTs have been well characterized in bacteria, yeast, and mammals compared to plants. The characterization of CPTs also has been less studied than TPTs. In the present study, molecular cloning and functional characterization of a CPT from a medicinal plant, Panax ginseng Mayer were addressed. Methods: Gene expression patterns of PgCPT1 were analyzed by quantitative reverse transcription polymerase chain reaction. In planta transformation was generated by floral dipping using Agrobacterium tumefaciens. Yeast transformation was performed by lithium acetate and heat-shock for $rer2{\Delta}$ complementation and yeast-two-hybrid assay. Results: The ginseng genome contains at least one family of three putative CPT genes. PgCPT1 is expressed in all organs, but more predominantly in the leaves. Overexpression of PgCPT1 did not show any plant growth defect, and its protein can complement yeast mutant $rer2{\Delta}$ via possible protein-protein interaction with PgCPTL2. Conclusion: Partial complementation of the yeast dolichol biosynthesis mutant $rer2{\Delta}$ suggested that PgCPT1 is involved in dolichol biosynthesis. Direct protein interaction between PgCPT1 and a human Nogo-B receptor homolog suggests that PgCPT1 requires an accessory component for proper function.

• Molecules and Cells
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• v.38 no.10
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• pp.876-885
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• 2015

N-acetyl-D-glucosamine kinase (NAGK) plays an enzyme activity-independent, non-canonical role in the dendritogenesis of hippocampal neurons in culture. In this study, we investigated its role in axonal development. We found NAGK was distributed throughout neurons until developmental stage 3 (axonal outgrowth), and that its axonal expression remarkably decreased during stage 4 (dendritic outgrowth) and became negligible in stage 5 (mature). Immunocytochemistry (ICC) showed colocalization of NAGK with tubulin in hippocampal neurons and with Golgi in somata, dendrites, and nascent axons. A proximity ligation assay (PLA) for NAGK and Golgi marker protein followed by ICC for tubulin or dynein light chain roadblock type 1 (DYNLRB1) in stage 3 neurons showed NAGK-Golgi complex colocalized with DYNLRB1 at the tips of microtubule (MT) fibers in axonal growth cones and in somatodendritic areas. PLAs for NAGK-dynein combined with tubulin or Golgi ICC showed similar signal patterns, indicating a three way interaction between NAGK, dynein, and Golgi in growing axons. In addition, overexpression of the NAGK gene and of kinase mutant NAGK genes increased axonal lengths, and knockdown of NAGK by small hairpin (sh) RNA reduced axonal lengths; suggesting a structural role for NAGK in axonal growth. Finally, transfection of 'DYNLRB1 (74-96)', a small peptide derived from DYNLRB1's C-terminal, which binds with NAGK, resulted in neurons with shorter axons in culture. The authors suggest a NAGK-dynein-Golgi tripartite interaction in growing axons is instrumental during early axonal development.

• Journal of Bio-Environment Control
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• v.11 no.3
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• pp.139-143
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• 2002

Cotton Glutathione S-Transferase (GST: EC 2.5.1.18) was cloned and overexpressed in tobacco (Nicotiana tabacum) plants. Northern blot analysis confirmed the successful transformation of cotton gst gene in tobacco plant. Type I and Type ll transcript patterns were identified in transgenic tobacco plants and only Type I transcripts were discussed in this paper, The activity of GST in the type II transgenic plants was about 1.5-fold higher than those of the wild type and non-expresser by using 1-chloro-2,4-dinitrobenzene (CDNB) and reduced glutathione as the substrate. The expression of cotton GST in tobacco plants proved that Gh-5 could be translated into functional protein. Type II transgenic plants produced functional GST in the cells. The effects of cotton GST in the seedlings was evaluated by growing the control and transgenic seedlings at $15^{\circ}C$ in the growth chamber in the light. Overexpressors were grown well compared to the control plants (non-expressors). lo test far tolerance to salinity, seeds of Gh-5 overexpressors and the wild type Xanthi seedlings were grown at 0, 50, 100, 150, and 200 mM NaCl solution. Gh-5 transgenic seedlings showed higher growth rate over control seedlings on 50 and 100 mM NaCl solution. There was no difference in growth rate at 150 and 200mM NaCl concentration.

• Journal of Veterinary Science
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• v.23 no.6
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• pp.90.01-90.13
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• 2022

Background: Insulin regulates glucose homeostasis and has important effects on metabolism, cell growth, and differentiation. Depending on the cell type and physiological context, insulin signal has specific pathways and biological outcomes in different tissues and cells. For studying the signal pathway of insulin on glycolipid metabolism in porcine embryonic fibroblast (PEF), we used high-throughput sequencing to monitor gene expression patterns regulated by insulin. Objectives: The goal of our research was to see how insulin affected glucose and lipid metabolism in PEFs. Methods: We cultured the PEFs with the addition of insulin and sampled them at 0, 48, and 72 h for RNA-Seq analysis in triplicate for each time point. Results: At 48 and 72 h, 801 and 1,176 genes were differentially expressed, respectively. Of these, 272 up-regulated genes and 264 down-regulated genes were common to both time points. Gene Ontology analysis was used to annotate the functions of the differentially expressed genes (DEGs), the biological processes related to lipid metabolism and cell cycle were dominant. And the DEGs were significantly enriched in interleukin-17 signaling pathway, phosphatidylinositol-3-kinase-protein kinase B signaling pathway, pyruvate metabolism, and others pathways related to lipid metabolism by Kyoto Encyclopedia of Genes and Genomes enrichment analysis. Conclusions: These results elucidate the transcriptomic response to insulin in PEF. The genes and pathways involved in the transcriptome mechanisms provide useful information for further research into the complicated molecular processes of insulin in PEF.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2001.06a
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• pp.40-45
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• 2001

Raw starch-digesting amylase (BF-2A, M.W. 93, 000 Da) from Bacillus circulans F-2 was converted to two components during digestion with subtilisin. Two components were separated and designated as BF-2A' (63, 000 Da) and BF-2B (30, 000 Da), respectively. BF-2A' exhibited the same hydrolysis curve for soluble starch as the original amylase (BF-2A). Moreover, the catalytic activities of original and modified enzymes were indistinguishable in $K_{m}$, Vmax for, and in their specific activity for soluble starch hydrolysis. However, its adsorbability and digestibility on raw starch was greatly decreased. Furthermore, the enzymatic action pattern on soluble starch was greatly different from that of the BF-2A. A smaller peptide (BF-2B) showed adsorb ability onto raw starch. By these results, it is suggested that the larger peptide (BF-2A') has a region responsible for the expression of the enzyme activity to hydrolyze soluble substrate, and the smaller peptide (BF-2B) plays a role on raw starch adsorption. A similar phenomenon is observed during limited proteinase K, thermolysin, and endopeptidase Glu-C proteolysis of the enzyme. Fragments resulting from proteolysis were characterized by immunoblotting with anti-RSDA. The proteolytic patterns resulting from proteinase K and subtilisin were the same, producing 63- and 30-kDa fragments. Similar patterns were obtained with endopeptidase Glu-C or thermolysin. All proteolytic digests contained a common, major 63-kDa fragment. Inactivation of RSDA activity results from splitting off the C-terminal domain. Hence, it seems probable that the protease sensitive locus is in a hinge region susceptible to cleavage. Extracellular enzymes immunoreactive toward anti-RSDA were detected through whole bacterial cultivation. Proteins of sizes 93-, 75-, 63-, 55-, 38-, and 31-kDa were immunologically identical to RSDA. Of these, the 75-kDa and 63-kDa proteins correspond to the major products of proteolysis with Glu-C and thermolysin. These results postulated that enzyme heterogeneity of the raw starch-hydrolysis system might arise from the endogeneous proteolytic activity of the bacterium. Truncated forms of rsda, in which the gene sequence encoding the conserved domain had been deleted, directed the synthesis of a functional amylase that did not bind to raw starch. This indicates that the conserved region of RSDA constitutes a raw starch-binding domain, which is distinct from the active centre. The possible role of this substrate-binding region is discussed.d.

• Journal of Society of Preventive Korean Medicine
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• v.21 no.2
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• pp.1-13
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• 2017

Objectives : This paper serves to explore current trends of systems biology in Traditional Chinese Medicine (TCM) and examine how it may influence the Traditional Korean medicine. Methods : Literature review method was collectively used to classify Introduction to systems biology, diagnosis and syndrome classification of systems biology in TCM perspective, physiotherapy including acupuncture, herbs and formula functions, TCM systems biology, and directions of academic development. Results : The term 'Systems biology' is coined as a combination of systems science and biology. It is a field of study that tries to understand living organism by establishing a theory based on an ideal model that analyzes and predicts the desired output with understanding of interrelationships and dynamics between variables. Systems biology has an integrated and multi-dimensional nature that observes the interaction among the elements constructing the network. The current state of systems biology in TCM is categorized into 4 parts: diagnosis and syndrome, physical therapy, herbs and formulas and academic development of TCM systems biology and its technology. Diagnosis and syndrome field is focusing on developing TCM into personalized medicine by clarifying Kidney yin deficiency patterns and metabolic differences among five patterns of diabetes and analyzing plasma metabolism and biomarkers of coronary heart disease patients. In the field of physical therapy such as acupuncture and moxibustion, researchers discovered the effect of stimulating acupoint ST40 on gene expression and the effects of acupuncture on treating functional dyspepsia and acute ischemic stroke. Herbs and formulas were analyzed with TCM network pharmacology. The therapeutic mechanisms of Si Wu Tang and its series formulas are explained by identifying potential active substances, targets and mechanism of action, including metabolic pathways of amino acid and fatty acid. For the academic development of TCM systems biology and its technology, it is necessary to integrate massive database, integrate pharmacokinetics and pharmacodynamics, as well as systems biology. It is also essential to establish a platform to maximize herbal treatment through accumulation of research data and diseases-specific, or drug-specific network combined with clinical experiences, and identify functions and roles of molecules in herbs and conduct animal-based studies within TCM frame. So far, few literature reviews exist for systems biology in traditional Korean medicine and they merely re-examine known efficacies of simple substances, herbs and formulas. For the future, it is necessary to identify specific mechanisms of working agents and targets to maximize the effects of traditional medicine modalities. Conclusions : Systems biology is widely accepted and studied in TCM and already advanced into a field known as 'TCM systems biology', which calls for the study of incorporating TCM and systems biology. It is time for traditional Korean medicine to acknowledge the importance of systems biology and present scientific basis of traditional medicine and establish the principles of diagnosis, prevention and treatment of diseases. By doing so, traditional Korean medicine would be innovated and further developed into a personalized medicine.

• Journal of Plant Biotechnology
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• v.35 no.4
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• pp.317-327
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• 2008

Phytocystatins, which are inhibitors of plant cysteine peptidases, are involved in the regulation of protein turnover and in the defense against insect pests and pathogens. Extensive searches in the Brassica rapa genome allowed the prediction of at least eight different phytocystatin genes on seven chromosomes in the B. rapa genome. Structure comparisons based on alignments of the all BrCYS ($\underline{B}$. $\underline{r}apa$ $phyto{\underline{cys}}tatin$) proteins using the CLUSTALW program revealed conservation of the three consensus motifs known to interact with the active site of cysteine peptidases. According to the phylogenetic analysis based on the deduced amino acid sequences, the eight BrCYS proteins were divided into several clusters related to the orthologous phytocystatin. The predicted three-dimensional structure models of the eight BrCYS proteins demonstrate that all of these proteins are similar to the reported crystal structure of oryzacystatin-I (OC-I). Digital northern and RT-PCR analyses indicated that the eight BrCYS genes exhibit different expression patterns in B. rapa tissues and respond differently to abiotic stimuli. The differences in gene structure and expression between the eight BrCYS genes suggest that these proteins may play diverse physiological roles in B. rapa and may interact with cysteine peptidases through different mechanisms.

• The Plant Pathology Journal
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• v.30 no.4
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• pp.343-354
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• 2014

Rice blast disease caused by Magnaporthe oryzae is one of the most serious diseases of cultivated rice (Oryza sativa L.) in most rice-growing regions of the world. In order to investigate early response genes in rice, we utilized the transcriptome analysis approach using a 300 K tilling microarray to rice leaves infected with compatible and incompatible M. oryzae strains. Prior to the microarray experiment, total RNA was validated by measuring the differential expression of rice defense-related marker genes (chitinase 2, barwin, PBZ1, and PR-10) by RT-PCR, and phytoalexins (sakuranetin and momilactone A) with HPLC. Microarray analysis revealed that 231 genes were up-regulated (>2 fold change, p < 0.05) in the incompatible interaction compared to the compatible one. Highly expressed genes were functionally characterized into metabolic processes and oxidation-reduction categories. The oxidative stress response was induced in both early and later infection stages. Biotic stress overview from MapMan analysis revealed that the phytohormone ethylene as well as signaling molecules jasmonic acid and salicylic acid is important for defense gene regulation. WRKY and Myb transcription factors were also involved in signal transduction processes. Additionally, receptor-like kinases were more likely associated with the defense response, and their expression patterns were validated by RT-PCR. Our results suggest that candidate genes, including receptor-like protein kinases, may play a key role in disease resistance against M. oryzae attack.

• Korean Journal of Plant Resources
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• v.30 no.5
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• pp.571-577
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• 2017

Abscisic acid (ABA) is an important phytohormone involved in abiotic stress tolerance in plants. The group A bZIP transcription factors play important roles in the ABA signaling pathway in Arabidopsis but little is known about their functions in rice. In our current study, we have isolated and characterized a group A bZIP transcription factor in rice, OsABF3 (Oryza sativa ABA responsive element binding factor 3). We examined the expression patterns of OsABF3 in various tissues and time course analysis after abiotic stress treatments such as drought, salinity, cold, oxidative stress, and ABA in rice. Subcellular localization analysis in maize protoplasts using a GFP fusion vector further indicated that OsABF3 is a nuclear protein. Moreover, in a yeast one-hybrid experiment, OsABF3 was shown to bind to ABA responsive elements (ABREs) and its N-terminal region found to be necessary to transactivate a downstream reporter. A homozygous T-DNA insertional mutant of OsABF3 is more sensitive to salinity, drought, and oxidative stress compared with wild type plants ＆ OsABF3OX plants. In addition, this Osabf3 mutant showed a significantly decreased sensitivity to high levels of ABA at germination and post-germination. Collectively, our present results indicate that OsABF3 functions as a transcriptional regulator that modulates the expression of abiotic stress-responsive genes through an ABA-dependent pathway.

• Journal of Food Hygiene and Safety
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• v.13 no.2
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• pp.171-187
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• 1998

Garlic occupies a special position among the many foods of vegetable origin because it is the sole food for Koreans during the their lives. And vitamin A has been ingested by forms of food or additives. Cadmium has been described as one of the most dangerous trace elements in the food and environment of man and livestocks. Since the de novo synthesis of stress proteins can be detected early after exposure to some agents, analysis of cadmium-induced changes in gene expression , ie. alterations in patterns of protein synthesis, may be useful to develop as biomarkers of exposure and damage for food hygiene. He acute and chronic combine effects of cadmium (Cd, CdCl2 20mg/kg), garlic oil(Dds: diallyl disulfide 50mg/kg, 3 times a week) and vitamin A(Ra: retinol acetate 50,000 IU/kg, 3 times a week) on Wistar male rats were evaluated concerning cadmium contents, tissues enzyme activity, HSP expression histopathological and electron microscopical examinations. The results of the study are as follows ; 1. Less cadmium was absorbed through the digestive tracts, but the ratio of contents in tissue were not changed by the simultaneous adminstration of diallyl disufide or retinol acetate. 2. ALT(alanine aminotransferase) , AST(aspartate aminotransferase), glucose, BUN (blood urea nitrogen), creatinine, the key indices of the clinical changes in hepatic and renal function were significantly hanged by the cadmium treatment after 1 week in liver, after 4 weeks in kidney. 3. Histopathological changes in cadmium treated rats were appeared at 8 weeks age treatment in kidneys. Homogenous eosinophilic material was accumulated in cortical and collecting tubular lumens at 16 weeks. Degenerated or necrotized tubular cells were observed in cortex and medulla. Degenerated seminiferous tubules and homogeneous eosinophilic material was seen in interstitial tissue of rat treated with cadmium for 16 weeks. Calcium deposits were seen in degenerated seminiferous tubules and the tubules showed severe calcification of rat treated with cadmium for 16 weeks. Electron microscope changes in kidney were observed in rats treated with CdCl2 20 mg/kg. Proximal convoluted tubule cells showed selling of cytoplasm and narrow lumen. Capillary endothelial cells showed cytoplasmic vacuoles and swelling. Degenerated epithelial cells were accumulated in tubular lumen of kidney. 4. Enhanced synthesis of 70 KDa relateve molecular mass proteins were detected in 2 hours after cadmium, exposure, with maximum activity occurring at 8~48 hours. Induction of HSP 70 was evident at proximal tubules and glomeruli in kidney. Testicular cells produced enough HSP to be detected normally. From the above results, it could be concluded that HSP70 induction by the cadmium treatment was a rapid reaction to indicated the exposure of xenobiotics, and retinol acetate reduced the cadmium induced nephrotoxicity.