• Journal of Ginseng Research
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• v.41 no.3
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• pp.419-427
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• 2017

Background: Glycosylation of natural compounds increases the diversity of secondary metabolites. Glycosylation steps are implicated not only in plant growth and development, but also in plant defense responses. Although the activities of uridine-dependent glycosyltransferases (UGTs) have long been recognized, and genes encoding them in several higher plants have been identified, the specific functions of UGTs in planta remain largely unknown. Methods: Spatial and temporal patterns of gene expression were analyzed by quantitative reverse transcription (qRT)-polymerase chain reaction (PCR) and GUS histochemical assay. In planta transformation in heterologous Arabidopsis was generated by floral dipping using Agrobacterium tumefaciens (C58C1). Protein localization was analyzed by confocal microscopy via fluorescent protein tagging. Results: PgUGT72AL1 was highly expressed in the rhizome, upper root, and youngest leaf compared with the other organs. GUS staining of the promoter: GUS fusion revealed high expression in different organs, including axillary leaf branch. Overexpression of PgUGT72AL1 resulted in a fused organ in the axillary leaf branch. Conclusion: PgUGT72AL1, which is phylogenetically close to PgUGT71A27, is involved in the production of ginsenoside compound K. Considering that compound K is not reported in raw ginseng material, further characterization of this gene may shed light on the biological function of ginsenosides in ginseng plant growth and development. The organ fusion phenotype could be caused by the defective growth of cells in the boundary region, commonly regulated by phytohormones such as auxins or brassinosteroids, and requires further analysis.

• Journal of Plant Biotechnology
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• v.50
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• pp.127-136
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• 2023

Water scarcity decreases the rate of photosynthesis and, consequently, the yield of banana plants (Musa spp). In this study, transcriptome analysis was performed to identify photosynthesis-related genes in banana plants and determine their expression profiles under water stress conditions. Banana plantlets were in vitro cultured on Murashige and Skoog agar medium with and without 10% polyethylene glycol and marked as BP10 and BK. Chlorophyll contents in the plant shoots were determined spectrophotometrically. Two cDNA libraries generated from BK and BP10 plantlets, respectively, were used as the reference for transcriptome data. Gene ontology (GO) enrichment analysis was performed using the Database for Annotation, Visualization, and Integrated Discovery (DAVID) and visualized using the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway prediction. Morphological observations indicated that water deficiency caused chlorosis and reduced the shoot chlorophyll content of banana plantlets. GO enrichment identified 52 photosynthesis-related genes that were affected by water stress. KEGG visualization revealed the pathways related to the 52 photosynthesisr-elated genes and their allocations in four GO terms. Four, 12, 15, and 21 genes were related to chlorophyll biosynthesis, the Calvin cycle, the photosynthetic electron transfer chain, and the light-harvesting complex, respectively. Differentially expressed gene (DEG) analysis using DESeq revealed that 45 genes were down-regulated, whereas seven genes were up-regulated. Four of the down-regulated genes were responsible for chlorophyll biosynthesis and appeared to cause the decrease in the banana leaf chlorophyll content. Among the annotated DEGs, MaPNDO, MaPSAL, and MaFEDA were selected and validated using quantitative real-time PCR.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.9
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• pp.1350-1357
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• 2017

Objective: The present study aimed to investigate whether the long-lasting, recurrent restricting of sows leads to the physiological and psychological reaction of discomfort. Methods: Sows (Large White) that had experienced restricting for about 0.5 or 3 years and agematched sows kept in a group housing system (loose sows) were compared. Pupillary light reflex parameters were measured at the weaning stage. Immediately after slaughter, blood samples were taken to measure serum cortisol levels, and the brain was dissected, gene expression in the hippo-campus, frontal cortex and hypothalamus was analyzed. Results: The serum cortisol levels were higher in the confined sows than in the loose sows. The full maturity, but not the young adolescent, confined sows had longer latency time in the onset of pupil constriction than their loose counterparts. Real-time polymerase chain reaction analyses revealed an increased expression of interleukin 6 mRNA in the hippocampus and decreased expression of brain derived neurotrophic factor mRNA in hippocampus and hypothalamus and to a lesser extent in the frontal cortex of the full maturity confined sows, compared with the full maturity loose sows. Conclusion: Taken together, these data indicated that recurrent restricting stress in full maturity sows leads to the physiological and psychological reaction of discomfort.

• BMB Reports
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• v.41 no.12
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• pp.868-874
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• 2008

Neurofilaments (NFs) are neuronal intermediate filaments composed of light (NF-L), middle (NF-M), and heavy (NF-H) subunits. NF-L self-assembles into a "core" filament with which NF-M or NF-H co-assembles to form the neuronal intermediate filament. Recent reports show that point mutations of the NF-L gene result in Charcot-Marie-Tooth disease (CMT). However, the most recently described rod domain mutant of human NF-L (A148V) has not been characterized in cellular level. We cloned human NF-L and used it to engineer the A148V. In phenotypic analysis using SW13 cells, A148V mutation completely abolished filament formation despite of presence of NF-M. Moreover, A148V mutation reduced the levels of in vitro self-assembly using GST-NF-L (H/R) fusion protein whereas control (A296T) mutant did not affect the filament formation. These results suggest that alanine at position 148 is essentially required for NF-L self-assembly leading to subsequent filament formation in neuronal cells.

• The Korean Journal of Mycology
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• v.22 no.1
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• pp.107-116
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• 1994

Interorder somatic hybrids were obtained by protoplast fusion between Pleurotus ostreatus in the order Agaricales and Elfvingia applanata in the order Aphyllophorales. The fusants were classified into stable heterokaryons and spontaneously segregated heterokaryons. Hyphae of all fusion products except two strains did not form clamp connections. Out of them, two clamped and three clampless fusants produced mature fruiting bodies by light-dark cycle on sawdust rice bran medium. All of these basidiocarps had clamp connections. Three fusants were analysed with the distribution of progenies and segregation of genetic characters by random spore analyses. The genetic markers were shown to segregate and recombine in the first generation of monospores isolated from basidiocarps. Phenotypes of a large number of auxotrophic progenies were not detected in the two clamped fusants. The aberration ratio of segregants indicated the gene interaction resulting from different genome structure between distantly related species. The polymerase chain reaction (PCR) was adopted for the detection of somatic hybrids nuclear DNA. Four fusants showed a positive results in three kinds of primers. The prominent reaction products are represented by new bands in primer # 87 and # 125. Out of four fusants, two somatic hybrids had non-parental mtDNA patterns when digested with EcoR1 and HindIII. Comparison of somatic hybrids, tissue culture isolates(TC) and multispore germination isolates(MS) were made using esterase isozyme analysis. It is apparent that somatic hybrids had a minor banding patterns which are quite different from those of parents.

• Clinical and Experimental Reproductive Medicine
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• v.43 no.1
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• pp.9-14
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• 2016

Objective: Autophagy contributes to the clearance and recycling of macromolecules and organelles in response to stress. We previously reported that vitrified mouse oocytes show acute increases in autophagy during warming. Herein, we investigate the potential role of Atg7 in oocyte vitrification by using an oocyte-specific deletion model of the Atg7 gene, a crucial upstream gene in the autophagic pathway. Methods: Oocyte-specific Atg7 deficient mice were generated by crossing Atg7 floxed mice and Zp3-Cre transgenic mice. The oocytes were vitrified-warmed and then subjected to in vitro fertilization and development. The rates of survival, fertilization, and development were assessed in the Atg7 deficient oocytes in comparison with the wildtype oocytes. Light chain 3 (LC3) immunofluorescence staining was performed to determine whether this method effectively evaluates the autophagy status of oocytes. Results: The survival rate of vitrified-warmed $Atg7^{f/f}$;Zp3-Cre ($Atg7^{d/d}$) metaphase II (MII) oocytes was not significantly different from that of the wildtype ($Atg7^{f/f}$) oocytes. Fertilization and development in the $Atg7^{d/d}$ oocytes were significantly lower than the $Atg7^{f/f}$ oocytes, comparable to the $Atg5^{d/d}$ oocytes previously described. Notably, the developmental rate improved slightly in vitrified-warmed $Atg7^{d/d}$ MII oocytes when compared to fresh $Atg7^{d/d}$ oocytes. LC3 immunofluorescence staining showed that this method can be reliably used to assess autophagic activation in oocytes. Conclusion: We confirmed that the LC3-positive signal is nearly absent in $Atg7^{d/d}$ oocytes. While autophagy is induced during the warming process after vitrification of MII oocytes, the Atg7 gene is not essential for survival of vitrified-warmed oocytes. Thus, induction of autophagy during warming of vitrified MII oocytes seems to be a natural response to manage cold or other cellular stresses.

• IMMUNE NETWORK
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• v.4 no.1
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• pp.7-15
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• 2004

Background: The heat shock proteins (HSPs) play an important role in cellular protection mechanisms against physical or chemical stresses. In this study scFv antibodies specific for human HSP70.1 were isolated from a semi-synthetic human scFv library with the ultimate goal of developing anti-HSP70.1 intracellular antibody (intrabody) that may offer an attractive alternative to gene targeting to study the function of the protein in cells. Methods: A semi-synthetic human scFv display library ($5{\times}10^{8}$ size) was constructed using pCANTAB-5E vector and the selection of the library against bacterially expressed recombinant human HSP70.1 was attempted by panning. Results: Three positive clones specific for recombinant HSP70.1 were identified. All three clones used $V_{H}$ subgroup III. On the other hand, $V_{L}$ of two clones belonged to the kappa light chain subgroup I, but the other utilized $V_{k}$ subgroup IV Interestingly, these scFv molecules specifically reacted to the recombinant HSP70.1, yet failed to recognize native HSP70 induced in U937 human monocytic cells by heat treatment. Conclusion: Our results indicated that affinity selection of an scFv phage display library using recombinant antigens produced in E. coli might not guarantee the isolation of scFv antibody molecules specific for a native form of the antigen. Therefore, the source of target antigens needs to be chosen carefully in order to isolate biofunctional antibody molecules.

• The Journal of the Korea Contents Association
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• v.9 no.4
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• pp.355-363
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• 2009

Increased oxidative stress by abnormal mitochondrial function can damage cell signal transduction and gene expression, and induce insulin resistance or diabetes. Autophagy, however, improve insulin resistance by clearance of malfunctioning mitochondria. Exercise also recovers the muscle dysfunction and degeneration by activating mitochondrial biogenesis. As it seems that exercise and autophagy might act as an orchestrated network to induce mitochondrial biogenesis, we investigated whether autophagy is involved in AMPK signal pathway stimulated by exercise or AICAR to increase mitochondrial biogenesis. And it showed that PGC-1 and mtTFA, but not autophagy marker LC3 mRNA expression were significantly increased by 6 hr of acute exercise. On the other hand, PGC-1 and mtTFA mRNA expression were upregulated by AICAR treatment to C2C12 myotube. However these genes were not inhibited by LC3 siRNA transfection. These results provide the evidence that autopahgy affects on mitochondrial biogenesis through different signal pathway from AMPK signal transduction.

• Journal of Life Science
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• v.21 no.5
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• pp.621-626
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• 2011

AMP-activated protein kinase (AMPK), a heterotrimeric complex comprising a catalytic ${\alpha}$ subunit and regulatory ${\beta}$ and ${\gamma}$ subunits, has been primarily studied as a major metabolic regulator in various organisms, but recent genetic studies discover its novel physiological functions. The first animal model with no functional AMPK ${\gamma}$ subunit gene was generated by using Drosophila genetics. AMPK ${\gamma}$ flies demonstrated lethality with severe defects in cuticle formation. Further histological analysis found that deletion of AMPK ${\gamma}$ causes severe defects in cell polarity in embryo epithelia. The phosphorylation of nonmuscle myosin regulatory light chain (MRLC), a critical regulator of epithelial cell polarity, was also diminished in AMPK ${\gamma}$ embryo epithelia. These defects in AMPK ${\gamma}$ mutant epithelia were successfully restored by over-expression of AMPK ${\gamma}$. Collectively, these results suggested that AMPK ${\gamma}$ is a critical cell polarity regulator in metazoan development.

• The Plant Pathology Journal
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• v.30 no.1
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• pp.58-67
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• 2014

The multifunctional triple gene block protein 1 (TGB1) of the Potexvirus Alternanthera mosaic virus (AltMV) has been reported to have silencing suppressor, cell-to-cell movement, and helicase functions. Yeast two hybrid screening using an Arabidopsis thaliana cDNA library with TGB1 as bait, and co-purification with TGB1 inclusion bodies identified several host proteins which interact with AltMV TGB1. Host protein interactions with TGB1 were confirmed by biomolecular fluorescence complementation, which showed positive TGB1 interaction with mitochondrial ATP synthase delta' chain subunit (ATP synthase delta'), light harvesting chlorophyll-protein complex I subunit A4 (LHCA4), chlorophyll a/b binding protein 1 (LHB1B2), chloroplast-localized IscA-like protein (ATCPISCA), and chloroplast ${\beta}$-ATPase. However, chloroplast ${\beta}$-ATPase interacts only with $TGB1_{L88}$, and not with weak silencing suppressor $TGB1_{L88}$. This selective interaction indicates that chloroplast ${\beta}$-ATPase is not required for AltMV movement and replication; however, TRV silencing of chloroplast ${\beta}$-ATPase in Nicotiana benthamiana induced severe tissue necrosis when plants were infected by AltMV $TGB1_{L88}$ but not AltMV $TGB1_{L88}$, suggesting that ${\beta}$-ATPase selectively responded to $TGB1_{L88}$ to induce defense responses.