• Molecules and Cells
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• v.38 no.3
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• pp.259-266
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• 2015

The regulation of flowering time has crucial implications for plant fitness. MicroRNA156 (miR156) represses the floral transition in Arabidopsis thaliana, but the mechanisms regulating its transcription remain unclear. Here, we show that two AGAMOUS-like proteins, AGL15 and AGL18, act as positive regulators of the expression of MIR156. Small RNA northern blot analysis revealed a significant decrease in the levels of mature miR156 in agl15 agl18 double mutants, but not in the single mutants, suggesting that AGL15 and AGL18 co-regulate miR156 expression. Histochemical analysis further indicated that the double mutants showed a reduction in MIR156 promoter strength. The double mutants also showed reduced abundance of pri-miR156a and pri-miR156c, two of the primary transcripts from MIR156 genes. Electrophoretic mobility shift assays demonstrated that AGL15 directly associated with the CArG motifs in the MIR156a/c promoters. AGL18 did not show binding affinity to the CArG motifs, but pull-down and yeast two-hybrid assays showed that AGL18 forms a heterodimer with AGL15. GFP reporter assays and bimolecular fluorescence complementation (BiFC) showed that AGL15 and AGL18 co-localize in the nucleus and confirmed their in vivo interaction. Overexpression of miR156 did not affect the levels of AGL15 and AGL18 transcripts. Taking these data together, we present a model for the transcriptional regulation of MIR156. In this model, AGL15 and AGL18 may form a complex along with other proteins, and bind to the CArG motifs of the promoters of MIR156 to activate the MIR156 expression.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.1
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• pp.367-372
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• 2015

Background: Human adiponectin (ApN), a 30 kDa glycoprotein of 244-amino acids which is predominantly produced by adipocytes, exerts its effects via two receptors, namely adiponectin receptor-1 (adipo-R1) and adiponectin receptor-2 (adipo-R2) with differential binding affinity to globular adiponectin. Adiponectin receptor expression has been studied in several cancer tissues. However, there are no studies of colorectal adenomas which are considered to be precursors for colorectal carcinoma (CRC). Objectives: In the present study, the expression of adipo-R1 and adipo-R2 was investigated immunohistochemically in colorectal adenomas and colorectal carcinoma tissues in an attempt to determine associations with these tumors. Materials and Methods: The study enrolled 50 CRC patients with tumor resection and 82 patients who were diagnosed with adenomatous polyps, classified as negative for neoplasia, low-grade dysplasia (L-GD) or high- grade dysplasia (H-GD). Results: Expression of both adipo-R1 and adipo-R2 was found to be significantly lower in the CRCs than in colorectal adenomas (tubular and tubulovillous, p=0.009 and p<0.001, respectively). Adipo-R1 and adipo-R2 expression was also significantly lower in the CRC group when compared with the groups of patients with low grade dysplasia, high-grade dysplasia or no neoplasia (p=0.012 and p<0.001, respectively). In addition, it was observed that adipo-R2 expression was generally positive in the non-neoplastic group irrespective of the adipo-R2 expression. In the L-GD, H-GD and CRC groups, the adipo-R2 result was positive whenever adipo-R1 result was positive but some patients with negative adipo-R1 had positive adipo-R2 (p<0.001, p=0.004, p<0.001, respectively). Conclusions: This study indicated that ApN may play a role in the progression of colorectal adenomatous polyps to carcinoma through actions on adipo-R1 and adipo-R2 receptors.

• Korean Journal of Microbiology
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• v.50 no.3
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• pp.239-244
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• 2014

ErmSF is one of the four antibiotic resistance factor proteins expressed by Streptomyces fradiae, antibiotic tylosin producer, which renders $MLS_B$ (macrolide-lincosamide-streptogramin B) antibiotic resistance through dimethylating A2058 of 23S rRNA, thereby reducing the affinity of antibiotic to ribosome. Unlike other Erm proteins, ErmSF harbors long N-terminal end region. To investigate its role in enzyme activity, mutant ErmSF deleted of 1-38 amino acids was overexpressed and activity in vivo and in vitro was observed. In vitro enzymatic assay showed that mutant protein exhibited reduced activity by 20% compared to the wild type enzyme. Due to the reduced activity of the mutant protein, cells expressing mutant protein showed weaker resistance to erythromycin than cells with wild type enzyme. Presumably, the decrease in enzyme activity was caused by the hindrance in substrate binding and (or) product release, not by defect in the methyl group transfer occurred in active site.

• Molecules and Cells
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• v.38 no.5
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• pp.409-415
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• 2015

Low-barrier hydrogen bonds (LBHBs) have been proposed to have important influences on the enormous reaction rate increases achieved by many enzymes. ${\Delta}^5$-3-ketosteroi isomerase (KSI) catalyzes the allylic isomerization of ${\Delta}^5$-3-ketosteroid to its conjugated ${\Delta}^4$-isomers at a rate that approache the diffusion limit. Tyr14, a catalytic residue of KSI, has been hypothesized to form an LBHB with the oxyanion of a dienolate steroid intermediate generated during the catalysis. The unusual chemical shift of a proton at 16.8 ppm in the nuclear magnetic resonance spectrum has been attributed to an LBHB between Tyr14 $O{\eta}$ and C3-O of equilenin an intermediate analogue, in the active site of D38N KSI. This shift in the spectrum was not observed in Y30F/Y55F/D38N and Y30F/Y55F/Y115F/D38N mutant KSIs when each mutant was complexed with equilenin, suggesting that Tyr14 could not form LBHB with the intermediate analogue in these mutant KSIs. The crystal structure of Y30F/Y55F/Y115F/D38N-equilenin complex revealed that the distance between Tyr14 $O{\eta}$ and C3-O of the bound steroi was within a direct hydrogen bond. The conversion of LBHB to an ordinary hydrogen bond in the mutant KSI reduced the binding affinity for the steroid inhibitors by a factor of 8.1-11. In addition, the absence of LBHB reduced the catalytic activity by only a factor of 1.7-2. These results suggest that the amount of stabilization energy of the reaction intermediate provided by LBHB is small compared with that provided by an ordinary hydrogen bond in KSI.

• Korean Journal of Veterinary Research
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• v.34 no.2
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• pp.307-313
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• 1994

We have previously shown that crude water extract of Euonymus alatus (EA) had strong prophylactic effect against chemically induced-and tumor cell implanted-cancer, and that the mechanisms responsible for its antitumor effects were due to nonspecific enhancement of the NK cell activities and the cell mediated immunity. However, it was unknown that any components of crude extract did work so, since it consisted of several components. In this paper, we fractionated the crude watar EA-extract into several fraction such as hexane-, ethylether-, ethyl acetate-, n-butanol- and water soluble-fraction, and screened the immune regulating activities of each fraction by the evaluation of lymphokine production and activated lymphocyte proliferation. As a result of the component fraction of EA-extract, it was found that n-butanol fraction was a potent immunostimulator, and the remained water soluble fraction also contained some stimulator, But, other fraction did not showed any remarkable effect. It is therefore suggested that EA-glycosides in n-butanol fraction may be new one of the potent biological response modifiers. The present study was also undertaken in an efforts to investigate the effects of elm-bark(EB, Ulmus clavidiana var japonica), which has been used for curing ulcer and inflammation as a folk medicine without any kind of experimental evidence to support this, on the cellular- and humoral-immune responses, lymphocyte function and NK cell activities in mice. Regardless of time and duration of EB-treatment, Arthus reaction and antibody response to SRBC were not modified by EB, but delayed hypersensitivity to SRBC was significantly enhanced only when EB was treated prior to SRBC-sensitization. EB slightly inhibited the proliferation responses of splenocytes to PHA-stimulation, but it significantly augmented the responses of these cells to S aureus Cowan 1 and Con A-activation, and these effects were manifested only when EB was added at culture initiation. EB did not influence Ig secretion of spleen cells but it significantly augmented the Con A-induced 1L 2 and MIF production of splenocytes. NK cell activities of splenocytes were markedly riled when effector cells were pretreated with EB and this augmentation was dine to the increase of binding affinity of effector cells to target cells and the target cell lytic activities of effector cells. These results led to the conclusion that EB triggers increase of cellular immune responses, such as delayed hypersensitivitiy, lymphokine production and NK cell activities. Also these results suggested that EB contains potent immune stimulants, which may provide the rational basis for their therapeutic use as one of the new biological response modifiers.

• Applied Microscopy
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• v.41 no.1
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• pp.37-45
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• 2011

Known as a female hormone, estrogen, has an effect on the male reproductive organs. The estrogen has to combine with the estrogen receptor to communicate a signal. Propyl pyrazole triol (PPT) is an estrogen receptor alpha selective agonist with a 410-, or 1,000-fold relative binding affinity for estrogen receptor alpha versus estrogen receptor beta. In this study, adult male mice were treated weekly with subcutaneously injections of PPT (0.01 mg, 0.1 mg, 1mg and 4 mg) suspended in castor oil (as control) for 8 weeks and observed histologically changes in testis, efferent ductule and epididymis. In the high concentrations of PPT 4 mg treatment group, a remarkable reduction was observed in the weight of the body, testis and epididymis. Microscopic examination revealed a reduction in seminiferous tubular diameter of the testis, and epithelial cell height of the epididymis in treated group during the experiment. In addition, as the diameter of the efferent ductule increased gradually, the height of epithelial cells was decreased. PPT 4 mg treatment group caused inhibition of spermatogenesis due to atrophied germinal epithelium in the testis, and decrease of adipocyte size attached to the epididymis. Sperm was not observed in the caudal epididymis of PPT 4 mg treated group. In conclusion, the injection of high concentrations of PPT into adult male mice induced physiological changes, such as an inhibition of spermatogenesis, and also histological changes within the reproductive organs.

• Korean Journal of Food Science and Technology
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• v.35 no.3
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• pp.379-385
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• 2003

Effects of molokhia (Corchorus olitorius) and its mucilage on in vitro bile acid adsorption capacity and lipid composition using cholesterol-fed SD rats were investigated. Mucilage showed stronger affinity toward bile acid than hot water extract of molokhia powder. As the extracting temperature increased from 50 to $80^{\circ}C$, bile acid binding capacity of mucilage also increased from 41 to 83%. When molokhia powder or its mucilage added into cholesterol diet at 5 or 10% levels (as fiber source) was compared with cellulose-added group, total cholesterol and triglyceride levels of plasma showed no significant differences, whereas, HDL-cholesterol level of cellulose group significantly increased. Accumulation of cholesterol and triglyceride in liver were significantly inhibited in molokhia and mucilage groups. Molokhia and mucilage lowered the liver weights significantly. As the concentration of molokhia or mucilage increased, cholesterol and triglyceride levels in the liver was lowered. Cholesterol and triglyceride excreted through feces were significant increased in molokhia- or mucilage-fed group, with excretion of cholesterol by molokhia-fed group being mere distinct.

• Food Science and Preservation
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• v.24 no.7
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• pp.885-890
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• 2017

For the construction of the microbial monitoring method, anti-Salmonella polyclonal antibodies (pAbs) were produced from a rabbit and purified by saturated ammonium sulfate precipitation and protein A affinity column. The reactivity of anti-Salmonella pAbs was compared to that of commercial ones by using an indirect ELISA. The specificity of anti-Salmonella pAbs was investigated using 20 Salmonella serotypes and 20 non-Salmonella strains. A capturing ability of anti-Salmonella pAbs was investigated by exposing antibody-immobilized gold biosensor to different concentration of Salmonella mixture. Anti-Salmonella pAbs were successfully produced and purified with an antibody concentration of 2.0 mg/mL The reactivity of purified anti-Salmonella pAbs was greater than that of commercial one at all tested concentrations. All Salmonella serotypes, except S. Diarizonae, showed excellent binding efficiency with purified anti-Salmonella pAbs. Moreover, the purified anti-Salmonella pAbs showed excellent specificity against all non-Salmonella strains. The anti-Salmonella pAbs immobilized on the gold biosensor demonstrated the successful capturing capability against Salmonella with a dose-response manner. Therefore, the anti-Salmonella pAbs exhibited sufficient reactivity, specificity, as well as capturing capability against Salmonella to be considered as a bio-recognition element.

• The Korean Journal of Physiology and Pharmacology
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• v.17 no.3
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• pp.223-228
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• 2013

The calcium-activated $K^+$ ($BK_{Ca}$) channel is one of the potassium-selective ion channels that are present in the nervous and vascular systems. $Ca^{2+}$ is the main regulator of $BK_{Ca}$ channel activation. The $BK_{Ca}$ channel contains two high affinity $Ca^{2+}$ binding sites, namely, regulators of $K^+$ conductance, RCK1 and the $Ca^{2+}$ bowl. Lysophosphatidic acid (LPA, 1-radyl-2-hydroxy-sn-glycero-3-phosphate) is one of the neurolipids. LPA affects diverse cellular functions on many cell types through G protein-coupled LPA receptor subtypes. The activation of LPA receptors induces transient elevation of intracellular $Ca^{2+}$ levels through diverse G proteins such as $G{\alpha}_{q/11}$, $G{\alpha}_i$, $G{\alpha}_{12/13}$, and $G{\alpha}s$ and the related signal transduction pathway. In the present study, we examined LPA effects on $BK_{Ca}$ channel activity expressed in Xenopus oocytes, which are known to endogenously express the LPA receptor. Treatment with LPA induced a large outward current in a reversible and concentration-dependent manner. However, repeated treatment with LPA induced a rapid desensitization, and the LPA receptor antagonist Ki16425 blocked LPA action. LPA-mediated $BK_{Ca}$ channel activation was also attenuated by the PLC inhibitor U-73122, $IP_3$ inhibitor 2-APB, $Ca^{2+}$ chelator BAPTA, or PKC inhibitor calphostin. In addition, mutations in RCK1 and RCK2 also attenuated LPA-mediated $BK_{Ca}$ channel activation. The present study indicates that LPA-mediated activation of the $BK_{Ca}$ channel is achieved through the PLC, $IP_3$, $Ca^{2+}$, and PKC pathway and that LPA-mediated activation of the $BK_{Ca}$ channel could be one of the biological effects of LPA in the nervous and vascular systems.

• Microbiology and Biotechnology Letters
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• v.39 no.3
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• pp.224-237
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• 2011

For establishing the efficacy against breast cancer occurrence, women's mammographic breast density and hot flush remission related to pre and post-menopausal symptoms were selected as biomarkers. Meta analysis applied the final selection of 40 papers from Medline to assess the efficacy of isoflavone-rich soy or supplements versus placebo under randomized controlled trials. Interestingly, the exposure to the purified isoflavones was more effective than the exposure to the crude extract of soy isoflavones for hot flush suppression. The length and the amount of dosage dependency is the most appropriately suggested to be one year long with 50~100 mg/day of isoflavone-rich soy or supplements. Funnel plots was used to interpret the results, overall effect of isoflavones on breast density in post-menopausal women was revealed to be less effect [effect size: 0.062, 95% confidence interval (CI): 0.005 to 0.12], than that of pre-menopausal women (effect size: 0.101, CI: -0.003 to 0.205). The reason why breast density was found higher among the pre-menopausal than the post-menopausal women is that the phytoestrogens of dietary isoflavones were antagonized against estrogen by the basis of estrogen receptor binding affinity. Overall, the Meta analysis reported that isoflavone had limited influence on breast density by suppressing the expansion by only 2%, while it was more effective to suppressing the hot flush, showing a dramatic decrease of 23%.