• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.97-97
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• 2017

Plant genomes include heterochromatic loci that consist of repetitive sequences and transposable elements. LTR retrotransposon is the major class of transposons in advanced plants in terms of proportion in plant genome. The elements contribute not only to genome size but also to genome stability and gene expression. A number of cases have been reported transposon insertions near genic regions affect crop traits such as fruit pigments, stress tolerance, and yields. Functional LTR retrotransposons produce extrachromosomal DNA from genomic RNA by reverse transcription that takes place within virus-like-particles (VLPs). DECREASED DNA METHYLATION 1 (DDM1) plays important roles in maintaining DNA methylation of heterochromatin affecting all sequence contexts, CG, CHG, and CHH. Previous studies showed that ddm1 mutant exhibits massive transcription of retrotransposons in Arabidopsis, but only few of them were able to create new insertions into the genome. RNA-dependent RNA POLYMERASE 6 (RDR6) is known to function in restricting accumulation of transposon RNA by processing the transcripts into 21-22 nt epigenetically activated small interfering RNA (easiRNA). We purified VLPs and sequence cDNA to identify functional LTR retrotransposons in Arabidopsis ddm1 and ddm1rdr6 plants. Over 20 LTR copia and gypsy families were detected in ddm1 and ddm1rdr6 sequencing libraries and most of them were not reported for mobility. In ddm1rdr6, short fragments of ATHILA gypsy elements were detected. It suggests easiRNAs might regulate reverse transcription steps. The highest enriched element among transposon loci was previously characterized EVADE element. It has been reported that active EVADE element is more efficiently silenced through female germline than male germline. By genetic analyses, we found ddm1 and rdr6 mutation affect maternal silencing of active EVADE elements. DDM1-GFP protein accumulated in megaspore mother cell but was not found in mature egg cell. The fusion protein was also found in early embryo and maternal DDM1-GFP allele was more dominantly expressed in the embryo. We observed localization of DDM1-GFP in Arabidopsis and DDM1-YFP in maize and found the proteins accumulated in dividing zone of root tips. Currently we are looking at cell cycle dependency of DDM1 expression using maize system. Among 10 AGO proteins in Arabidopsis, AGO9 is specifically expressed in egg cell and shoot meristematic cells. In addition, mutation of AGO9 and RDR6 caused failure in maternal silencing, implying 21-22 nt easiRNA pathway is important for retrotransposon silencing in female gametophyte or/and early embryo. On the other hand, canonical 24 nt sRNA-directed DNA methylation (RdDM) pathways did not contribute to maternal silencing as confirmed by this study. Heat-activated LTR retrotransposon, ONSEN, was not silenced by DDM1 but the silencing mechanisms require RdDM pathways in somatic cells. We will propose distinct mechanisms of LTR retrotransposons in germline and somatic stages.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.13
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• pp.5437-5442
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• 2014

Esophageal squamous cell carcinoma (ESCC) is one of the most malignancies with a poor prognosis. The phospholipase $C{\varepsilon}$ gene (PLCE1) encodes a novel ras-related protein effector mediating the effects of R-Ras on the actin cytoskeleton and membrane protrusion. However, molecular mechanisms pertinent to ESCC are unclear. We therefore designed PLCE1-special small interfering RNA and transfected to esophageal squamous cell (EC) 9706 cells to investigat the effects of PLCE1 gene silencing on the cell cycle and apoptosis of ESCC and indicate its important role in the development of ESCC. Esophageal cancer tissue specimens and normal esophageal mucosa were obtained and assayed by immunohistochemical staining to confirm overexpression of PLCE1 in neoplasias. Fluorescence microscopy was used to examine transfection efficiency, while the result of PLCE1 silencing was examined by reverse transcription (RT-PCR). Flow cytometry and annexin V apoptosis assays were used to assess the cell cycle and apoptosis, respectively. Expression of cyclin D1 and caspase-3 was detected by Western-blotting. The level of PLCE1 protein in esophageal cancer tissue was significantly higher than that in normal tissue. After transfection, the expression of PLCE1 mRNA in EC 9706 was significantly reduced, compared with the control group. Furthermore, flow cytometry results suggested that the PLCE1 gene silencing arrested the cell cycle in the G0/G1 phase; apoptosis was significantly higher than in the negative control group and mock group. PLCE1 gene silencing by RNAi resulted in decreased expression of cyclin D1 and increased expression of caspase-3. Our study suggests that PLCE1 may be an oncogene and play an important role in esophageal carcinogenesis through regulating proteins which control cell cycling and apoptosis.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.5
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• pp.670-677
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• 2004

Sixteen multiparous Holstein-Friesian crossbred cows in mid-lactation were blocked according to days in milk (DIM) and previous lactation and randomly assigned according to a Randomized Complete Block (RCB) design with four replications to receive four dietary treatments. The dietary treatments consisted of T1: No cassava hay (CH) or stylo 184 hay (SH) supplementation, supplementation of concentrate to milk yield at 1:2 (control), T2: Supplementation of 1 kg of CH/hd/d, supplementation of concentrate to milk yield at 1:2, T3: Supplementation of 1 kg of CH+SH/hd/d, supplementation of concentrate to milk yield at 1:2, T4: Supplementation of 2 kg of CH+SH/hd/d, supplementation of concentrate to milk yield at 1:3. All animals received Ruzi grass from a cut-and-carry system as roughage source. The feeding trial lasted for 9 weeks. The results revealed that DMI of concentrate of supplemented treatments were significantly lower (p<0.05) than those in the control, but there was no significant difference between T2 and T3. There was no significant difference in forage DM intake between the control and supplemented treatments. CP and NDF digestibility of supplemented treatments were significantly (p<0.05) greater than the control and there were no significant differences among supplemented treatments. Milk yield and 3.5% FCM (14.3, 14.5, 14.7 and 14.8; 13.9, 14.3, 14.3 and 14.6 kg/hd/d, respectively) were not significantly different among treatments. Milk protein percentage of supplemented treatments was significantly (p<0.05) higher than the control, but there were no significant differences among supplemented treatments. There was no significant difference in milk fat percentage between the control and supplemented treatments. However, milk fat percentage tended to be higher for supplemented animals as compared to the control group. There were also no significant differences in lactose, solids-not-fat and total solids percentages among treatments. Cows in supplemented treatments gave incomes over supplement cost (IOSC) of 2.72, 2.74 and 2.93 US$/hd/d, respectively which were greater than for cows on control treatment. Furthermore, IOSC were greatest for cows in T4 as compared to other treatments. Based on this study it was concluded that, feeding cassava hay solely or in combination with stylo 184 hay as a supplemental protein source could be a potential valuable strategy in small-holder dairy farming systems in the tropics. This strategic supplementation significantly reduced concentrate use, which resulted in improved milk yields and milk quality for the supplemented cows. Moreover, it resulted in higher economical returns through increased productivity and lower ratios of concentrate to milk yield, from 1:2 to 1:3.

• Asian-Australasian Journal of Animal Sciences
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• v.15 no.5
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• pp.650-657
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• 2002

Two experiments were conducted to evaluate the effects of forage:concentrate ratios and feeding principles on milk yield, milk composition, body weight change, postpartum oestrus and feed cost. A total of 36 crossbred F1 cows (Holstein Friesian${\times}$Local Yellow) in the 8th week of lactation were used. In each experiment, animals were divided into three groups using a randomized block design according to the milk yield of the previous eight weeks. Cows were fed 30, 50 and 70% concentrate in the diet based on DM. In experiment 1 (Fc), cows were given the same amount of DM with constant ratios of forage and concentrate within treatments. In experiment 2 (Fa), cows were given the same constant amounts of concentrate as in experiment 1 and ad libitum forage. The forage consisted of a natural grass mixture based on 5 species of grasses with high nutritive values. There was no difference in total DM intake between treatments within experiments. However, cows fed forage ad libitum had higher DM intakes compared to cows fed constant forage (1.6, 4.5 and 9.5% for cows fed 70, 50 and 30% forage, respectively). Daily milk yield of cows fed forage ad libitum was higher than that of cows fed constant forage:concentrate ratio. Within experiment, milk yield was highest for cows fed 30% DM forage, followed by cows fed 50% and then cows fed 70% forage (11.17, 10.98 and 10.71 for the 30Fc, 50Fc and 70Fc cows; 11.73, 11.16 and 10.81 kg for the 30Fa, 50Fa and 70Fa cows, respectively). Decreased forage ratio in the diets resulted in decreased milk fat content and tended to increase milk protein. Increased concentrate ratio in the diet and feeding forage ad libitum increased body weight gain. The effect of forage:concentrate ratio on postpartum oestrus was not significant. The feed cost per kg milk produced was lowest for the cows fed 70% forage. It is concluded that increased ratio of concentrate resulted in increased body weight gain, milk yield, milk protein, and decreased milk fat. Feeding forage ad libitum increased feed intake, milk yield and body weight gain. The ratio of 50% forage is more suitable for milk production and animal condition, but in terms of feed cost and under the conditions of small dairy farmers, the 70% ad libitum forage feeding is recommended.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.24
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• pp.10879-10882
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• 2015

Objective: To investigate the effect of intraoperative glucose fluctuation and postoperative interlukin-6 (IL-6), tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), C-reactive protein (CRP) levels on the short-term prognosis of patients with intracranial supratentorial neoplasms. Materials and Methods: Eighty-six patients undergoing intracranial excision were selected in The Second Hospital of Jilin University. According to the condition of glucose fluctuation, the patients were divided into group A (glucose fluctuation <2.2 mmol/L, n=57) and group B (glucose fluctuation ${\geq}2.2mmol/L$, n=29). Glucose was assessed by drawing 2 mL blood from internal jugular vein in two groups in the following time points, namely fasting blood glucose 1 d before operation ($T_0$), 5 min after anesthesia induction ($T_1$), intraoperative peak glucose ($T_2$), intraoperative lowest glucose ($T_3$), 5 min after closing the skull ($T_4$), immediately after returning to intensive care unit (ICU) ($T_5$) and 2 h after returning to ICU ($T_6$). 1 d before operation and 1, 3 and 6 d after operation, serum IL-6 and TNF-${\alpha}$ levels were detected with enzyme-linked immunosorbent assay (ELISA), and CRP level with immunoturbidimetry. Additionally, postoperative adverse reactions were monitored. Results: There was no statistical significance between two groups regarding the operation time, anesthesia time, amount of intraoperative bleeding and blood transfusion (P>0.05). The glucose levels in both groups at $T_1{\sim}T_6$ went up conspicuously compared with that at $T_0$ (P<0.01), and those in group B at $T_2$, $T_4$, $T_5$ and $T_6$ were significantly higher than in group A (P<0.01). Serum IL-6, TNF-${\alpha}$ and CRP levels in both groups 1, 3 and 6 d after operation increased markedly compared with 1 d before operation (P<0.01), but the increased range in group A was notably lower than in group B (P<0.05 or P<0.01). Postoperative incidences of hypoglycemia, hyperglycemia and myocardial ischemia in group A were significantly lower than in group B (P<0.05), and respiratory support time obviously shorter than in group B (P<0.01). Conclusions: The glucose fluctuation of patients undergoing intracranial excision is related to postoperative IL-6, TNF-${\alpha}$ and CRP levels and those with small range of glucose fluctuation have better prognosis.

• BMB Reports
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• v.37 no.6
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• pp.720-725
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• 2004

A number of signaling molecules contain small pleckstrin homology (PH) domains capable of binding phosphoinositides or proteins. Phospholipase C (PLC)-${\gamma}1$ has two putative PH domains, an $NH_2$-terminal (PH1) and a split PH domain ($nPH_2$ and $cPH_2$). We previously reported that the split PH domain of PLC-${\gamma}1$ binds to phosphatidylinositol 4-phosphate (PI(4)P) and phosphatidylinositol 4,5-bisphosphate (PI(4,5)$P_2$) (Chang et al., 2002). To identify the amino acid residues responsible for binding with PI(4)P and PI(4,5)$P_2$, we used site-directed mutagenesis to replace each amino acid in the variable loop-1 (VL-1) region of the PLC-${\gamma}1$ $nPH_2$ domain with alanine (a neutral amino acid). The phosphoinositide-binding affinity of these mutant molecules was analyzed by Dot-blot assay followed by ECL detection. We found that two PLC-${\gamma}1$ nPH2 domain mutants, P500A and H503A, showed reduced affinities for phosphoinositide binding. Furthermore, these mutant PLC-${\gamma}1$ molecules showed reduced PI(4,5)$P_2$ hydrolysis. Using green fluorescent protein (GFP) fusion protein system, we showed that both $PH_1$ and $nPH_2$ domains are responsible for membrane-targeted translocation of PLC-${\gamma}1$ upon serum stimulation. Together, our data reveal that the amino acid residues $Pro^{500}$ and $His^{503}$ are critical for binding of PLC-${\gamma}1$ to one of its substrates, PI(4,5)$P_2$ in the membrane.

• Korean Journal of Microbiology
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• v.50 no.1
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• pp.73-83
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• 2014

The objective of the this study was to investigate the binding capacity and removal ability of lactic acid bacterial strains obtained from Korean soybean paste for mutagenic heterocyclic amines (HCAs) formed during cooking of protein-rich food at high temperature. Among 19 strains identified by carbohydrate fermentation and 16S rRNA sequencing, the live cell or cell-free culture supernatant of Lactobacillus acidophilus D11, Enterococcus faecium D12, Pediococcus acidilactici D19, L. acidophilus D38, Lactobacillus sakei D44, Enterococcus faecalis D66, and Lactobacillus plantarum D70 inhibited the mutagenesis caused by either 3-amino-1,4-dimethyl-5H-pyrido[4,3-b] indole (Trp-P-1) or 3-amino-1-methyl-5H-pyrido[4,3-b] indole (Trp-P-2) in Salmonella typhimurium TA98 and TA100. The bacterial cells of the isolated strains showed greater binding activity than the pure cell wall, exopolysaccharide, and pepetidoglycan. The carbohydrate moieties of the cell wall or protein molecules on the cell surface have a significant role in binding Trp-P-1 and Trp-P-2, since protease, heating, sodium metaperiodate, or acidic pH treatments significantly (P<0.05) reduced the binding efficacy of the tested bacteria. Addition of metal ions or sodium dodecyl sulfate decreased the binding ability of E. faecium D12, L. acidophilus D38, and E. faecalis D66. Therefore, the binding mechanisms of these strains may consist of ion-exchange and hydrophobic bonds. Especially, the high mutagen binding by L. acidophilus D38 and L. plantarum D70 may reduce the accumulation or absorption of Trp-P-1 and Trp-P-2 in the small intestine via increased excretion of a mutagen-bacteria complex.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.39 no.3
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• pp.261-268
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• 2006

Fish-frames are processing byproducts, which are left after obtaining fillets or muscle during fish processing. The fish-frame generally consists of muscle, collagen, calcium, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). We used fish-frame powder (FFP) of chum salmon and skipjack tuna to prepare and characterize snacks for human consumption with different proportions of FFP. The crude protein and lipid contents of fish-frames were 16.3 and 9.4% for chum salmon and 18.6 and 8.3% for skipjack tuna, respectively. The volatile basic nitrogen (30.6 mg/100 g) and browning index (0.393) of FFP from chum salmon were lower than those of FFP from skipjack tuna. Thus, the FFP of chum salmon was better for making snacks than that of skipjack tuna. Five snacks were prepared with 0, 10, 20, 30, and 40% (w/w) substitution ratios of FFP from chum salmon. The moisture content of the snacks decreased (33.6 to 11.5%) with increasing FFP substitution ratio, whereas crude ash (2.9 to 7.5%), protein (11.4 to 18.4%) and lipid (13.7 to 35.1%) increased. Sensory scores for the texture and taste of the snack with 30% FFP were significantly higher (p<0.05) than those for other snacks; the color and flavor scores of all snacks did not differ significantly. The major fatty acids in the snacks were 16:0 and 18:0 as saturates, 18:1n-9 as monoenes, and 18:2n-6 and 18:3n-3 as polyenes. Snacks with FFP contained small amounts of EPA (0.5 to 0.8%) and DHA (1.3 to 1.8%) in the total lipid composition. The total amino acid content (16.08 g/100 g) of the snack with 30% FFP was higher than that of the snack without FFP (11.18 g/100 g), and the major amino acids were aspartic acid, glutamic acid, glycine, leucine, and lysine. The calcium and phosphorus contents of the snack with 30% FFP were 1,272 mg/100 g and 854 mg/100 g, respectively, and their ratio was the optimal range (2:1 to 1:2) for body absorption efficiency.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1995.06b
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• pp.27-53
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• 1995

In plant pathology there is an increasing necessity for improved cytological techniques as basis for the localization of cellular substances within the dynamic fine structure of the host-(plant)-pathogen-interaction. Low temperature (LT) preparation techniques (shock freezing, freeze substitution, LT embedding) are now successfully applied in plant pathology. They are regarded as important tools to stabilize the dynamic plant-pathogen-interaction as it exists under physiological conditions. - The main advantage of LT techniques versus conventional chemical fixation is seen in the maintenance of the hydration shell of molecules and macromolecular structures. This results in an improved fine structural preservation and in a superior retention of the antigenicity of proteins. - A well defined ultrastructure of small, fungal organisms and large biological samples such as plant material and as well as the plant-pathogen (fungus) infection sites are presented. The mesophyll tissue of Arabidopsis thaliana is characterized by homogeneously structured cytoplasm closely attached to the cell wall. From analyses of the compatible interaction between Erysiphe graminis f. sp. hordei on barley (Hordeum vulgare), various steps in the infection sequence can be identified. Infection sites of powdery mildew on primary leaves of barley are analysed with regard to the fine structural preservation of the haustoria. The presentation s focussed on the ultrastructure of the extrahaustorial matrix and the extrahaustorial membrane. - The integration of improved cellular preservation with a molecular analysis of the infected host cell is achieved by the application of secondary probing techniques, i.e. immunocytochemistry. Recent data on the characterization of freeze substituted powdery mildew and urst infected plant tissue by immunogold methodology are described with special emphasis on the localization of THRGP-like (threonine-hydrxyproline-rich glycoprotein) epitopes. Infection sites of powdery mildew on barley, stem rust as well as leaf rust (Puccinia recondita) on primary leaves of wheat were probed with a polyclonal antiserum to maize THRGP. Cross-reactivity with the anti-THRGP antiserum was observed over the extrahaustorial matrix of the both compatible and incompatible plant-pathogen interactions. The highly localized accumulation of THRGP-like epitopes at the extrahaustorial host-pathogen interface suggests the involvement of structural, interfacial proteins during the infection of monocotyledonous plants by obligate, biotrophic fungi.

• Journal of The Korean Society of Grassland and Forage Science
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• v.33 no.3
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• pp.159-166
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• 2013

We have previously investigated the proteome changes of rice leaves under heat stress (Lee et al. in Proteomics 2007a, 7:3369-3383), wherein a group of antioxidant proteins and heat shock proteins (HSPs) were found to be regulated differently. The present study focuses on the biochemical changes and gene expression profiles of heat shock protein and antioxidant genes in rice leaves in response to heat stress ($42^{\circ}C$) during a wide range of exposure times. The results show that hydrogen peroxide and proline contents increased significantly, suggesting an oxidative burst and osmotic imbalance under heat stress. The mRNA levels of chaperone 60, HSP70, HSP100, chloroplastic HSP26, and mitochondrial small HSP responded rapidly and showed maximum expression after 0.5 or 2 h under heat stress. Transcript levels of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR) and Cu-Zn superoxide dismutase (Cu-Zn SOD) showed a rapid and marked accumulation upon heat stress. While prolonged exposure to heat stress resulted in increased transcript levels of monodehydroascorbate reductase, peroxidase, glyoxalase 1, glutathione reductase, thioredoxin peroxidase, 2-Cysteine peroxiredoxin, and nucleoside diphosphate kinase 1, while the transcription of catalase was suppressed. Consistent with their changes in gene expression, the enzyme activities of APX and DHAR also increased significantly following exposure to heat stress. These results suggest that oxidative stress is usually caused by heat stress, and plants apply complex HSP- and antioxidant-mediated defense mechanisms to cope with heat stress.