• The Korean Journal of Mycology
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• v.42 no.3
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• pp.191-200
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• 2014

Nuruk samples collected from various regions in Korea were investigated in terms of fungal contents and diversity. In measurement of colony forming unit (CFU) in Nuruk suspensions on DRBC agar, Nuruk samples MS4, MS8, and MS10 were among the highest fungal density, with $1,278.9{\pm}21.6$ (${\times}10^4$), $1,868.0{\pm}27.7$ (${\times}10^4$), and $775.1{\pm}19.2$ (${\times}10^4$) were among the samples showing the highest fungal density. CFU per 20 mg Nuruk, respectively. The majority of fungal components were yeasts, including Pichia anomala, P. kudriavzevii, Kluyveromyces marxianus, and Saccharomycopsis fibuligera, whereas Aspergillus oryzae and Rhizopus oryzae, the representative Nuruk fungi, were predominant only in the low fungal density Nuruks (MS2, MS5, and MS11). Saccharification capability of the fungal isolates was assessed by measurement of amylase activity in the culture broth. The highest amylase activity was found in A. niger and A. luchuensis, followed by S. fibuligera. A. oryzae and R. oryzae showed fair amylase activity but significantly lower than those of the three fungal species. R. oryzae was suggested to play an additional role in degradation of ${\beta}$-glucan in crop component of Nuruk since R. oryzae was the only fungus that showed ${\beta}$-glucanase activity among the fungal isolates. To confirm the safety of Nuruk, aflatoxigenicity of the isolated Aspergillus was estimated using the DNA markers norB-cypA, aflR, and omtA. All of the isolates turned out to be non-aflatoxigenic as evidenced by the deletion of gene markers, norB-cypA and aflR, and the absence of aflatoxin in the culture supernatants shown by TLC analysis.

• Korean Journal of Weed Science
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• v.30 no.2
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• pp.122-134
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• 2010

We investigated the levels of resistance and accumulation of terapyrroles, reactive oxygen species, lipid peroxidation, and antioxidative enzymes for reasons of growth reduction in herbicide-transgenic rice overexpressing Myxococcus xanthus, Arabidopsis thaliana, and human protoporphyrinogen oxidase (Protox) genes. The transgenic rice overexpressing M. xanthus (MX, MX1, PX), A. thaliana (AP31, AP36, AP37), and human (H45, H48, H49) Protox genes showed 43~65, 41~72 and 17~70-fold more resistance to oxyfluorfen, respectively, than the wild type. Among transgenic rice lines overexpressing Protox genes, several lines showed normal growth compared with the wild type, but several lines showed in reduction of plant height and shoot fresh weight under different light conditions. However, reduction of plant height of AP37 was much higher than other lines for the experimental period. On the other hand, the reduction of plant height and shoot fresh weight in the transgenic rice was higher in high light condition than in low light condition. Enhanced levels of Proto IX were observed in transgenic lines AP31, AP37, and H48 at 7 days after seeding (DAS) and transgenic lines PX, AP37, and H48 at 14 DAS relative to wild type. There were no differences in Mg-Proto IX of transgenic lines except for H41 and H48 and Mg-Proto IX monomethyl ester of transgenic lines except for MX, MX1, and PX. Although accumulation of tetrapyrrole intermediates was observed in transgenic lines, their tetrapyrrole accumulation levels were not enough to inhibit growth of transgenic rice. There were no differences in reactive oxygen species, MDA, ALA synthesizing capacity, and chlorophyll between transgenic lines and wild type indicating that accumulated tetrapyrrole intermediate were apparently not high enough to inhibit growth of transgenic rice. Therefore, the growth reduction in certain transgenic lines may not be caused by a single factor such as Proto IX, but by interaction of many other factors.

• Journal of Life Science
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• v.25 no.6
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• pp.680-685
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• 2015

Previously, cellulase and xylanase producing microorganism, Bacillus subtilis NC1, was isolated from soil. Based on the 16S rRNA gene sequence and API 50 CHL test the strain was identified as Bacillus subtilis, and named as B. subtilis NC1. We cloned and sequenced the genes for cellulase and xylanase. Plus, the deduced amino acid sequences from the genes of cellulase and xylanase were determined and were also identified as glycosyl hydrolases family (GH) 5 and 30, respectively. In this study to optimize the medium parameters for cellulase production by B. subtilis NC1 the RSM (response surface methodology) based on CCD (central composite design) model was performed. Three factors, tryptone, yeast extract, and NaCl, for N or C source were investigated. The cellulase activity was measured with a carboxylmethyl cellulose (CMC) plate and the 3,5-dinitrosalicylic acid (DNS) methods. The coefficient of determination (R²) for the model was 0.960, and the probability value (p=0.0001) of the regression model was highly significant. Based on the RSM, the optimum conditions for cellulase production by B. subtilis NC1 were predicted to be tryptone of 2.5%, yeast extract of 0.5%, and NaCl of 1.0%. Through the model verification, cellulase activity of Bacillus subtilis NC1 increased from 0.5 to 0.62 U/ml (24%) compared to the original medium.

• Journal of Life Science
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• v.19 no.11
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• pp.1637-1643
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• 2009

Shikonin, a component of Lithospermum erythrorhizon Sieb. et Zucc, exerts various characteristics such as anti-inflammatory, anti-cancer and anti-obesity functions. To elucidate the molecular mechanism of shikonin-induced inhibition of adipogenesis, we analyzed the mRNA expression level of various adipogenesis-related factors including C/EBPs (CCAAT/enhancerbinding proteins) and $PPAR{\gamma}$ (peroxisome proliferator-activated receptor $\gamma$). The data showed that mRNA expressions of C/$EBP{\beta}$ and C/$EPB{\delta}$ were only slightly changed by shikonin treatment, but mRNA expressions of $PPAR{\gamma}$ and C/$EPB{\alpha}$ were significantly down-regulated. Then, we tested whether upstream regulators of C/$EBP{\beta}$ and $PPAR{\gamma}$ were involved in anti-adipogenesis of shikonin. C/$EBP{\gamma}$ and CHOP (C/EBP homologous protein), which are upstream regulators of C/$EBP{\beta}$, were not affected by shikonin treatment. On the contrary, the mRNA level of KROX20 was markedly down-regulated by shikonin treatment. These results suggest that KROX20 might regulate downstream factors of adipogenesis through C/$EBP{\beta}$-independent pathway. The expression of KLF15 (Kruppel-like factor15), which is a member of KLF family and is a upstream regulator of $PPAR{\gamma}$, was dramatically decreased by shikonin treatment, but KLF2 was not changed. Shikonin had no impact on the expression of KLF5 in the early stage of adipogenesis, but shikonin increased expression of KLF5 in the late stage of adipogenesis. Even though mRNA expression of KLF5 was moderately changed by shikonin treatment, its effect may be small compared with the effect of KLF15, which was markedly inhibited. Taken together, these results suggest that shikonin inhibits adipogenesis through the down-regulation of $PPAR{\gamma}$ and C/$EPB{\alpha}$, which is mediated by the down-regulation of two pro-adipogenic factors, KROX20 and KLF15.

• Journal of Life Science
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• v.20 no.3
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• pp.381-387
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• 2010

To find offspring of Jeju Black cattle (JBC) produced by embryo transfer (ET) and artificial insemination (AI), a molecular genetic study was carried out in candidate cattle populations collected from cattle farms in Jeju Island, Korea. The genetic marker set was composed of 11 ISAG microsatellite (MS) markers, 11 SAES MS markers selected by our preliminary analysis for population diversity of JBC and two major coat color related genes: MC1R and ASIP. The results showed a combined non-exclusion probability for first parent (NE-P1) that was higher than that recommended by ISAG (above 0.9995), and a combined non-exclusion probability for sib identity of $5.3{\times}10^{-10}$. Parentage analysis showed that the cases identified the candidate's father only (77.0%), mother only (54.0%), and both parents (40.5%) in the candidate offspring population. The ET and AI calves were identified as 14.7% in the in vitro fertilized eggs provided and 32.4% in total population, respectively. However, the result from ISAG marker analysis showed 3 identical allele-combinations in 7 calves, and that from ISAG/SAES MS marker combination also showed 1 identical allele-combination in 2 calves. Data from MS and coat-color gene analyses provided information for complete identification of all animals tested. Because the present JBC population was mostly bred using small nuclear founders through bioengineering techniques such as AI and ET, the genetic diversity levels obtained from MS analysis in the JBC population were relatively lower than those of other cattle populations, including Hanwoo. The results suggested that the more efficient marker combinations, including coat color related genotypes, should be studied and used for constructing a system for identification and molecular breeding of JBC as well.

• Journal of Life Science
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• v.19 no.8
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• pp.1039-1046
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• 2009

The cytochrome P450s (P450s) metabolizing natural products are among the most versatile biological catalysts known in plants, but knowledge of the structural basis for their broad substrate specificity has been limited. The activity of p-coumarate 3-hydroxylase (C3H) is thought to be essential for the biosynthesis of lignin and many other phenylpropanoid pathway products in plants however, all attempts to express and purify the protein corresponding C3H gene have failed. As a result, no conditions suitable for the unambiguous assay of the enzyme are known. The detailed understanding of the mechanism and substrate-specificity of C3Hdemands a method for the production of active protein on the milligram scale. We have developed a bacterial expression and purification system for the plant C3H, which allows for the quick expression and purification of active wild-type C3H via introduction of combinational mutagenesis. The modified cytochrome P450 C3H ($C3H_{mod}$) could be purified in the absence of detergent using immobilized metal affinity chromatography and size exclusion chromatography following extraction from isolated membranes in a high salt buffer and catalytically activated. This method makes the use of isotopic labeling of C3H for NMRstudies and X-ray crystallography practical, and is also applicable to other plant cytochrome P450 proteins.

• Journal of Life Science
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• v.24 no.8
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• pp.820-826
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• 2014

The localization to specific subcellular sites and the regulation of cell surface receptors and channels are crucial for proper functioning. Postsynaptic density-95/Disks large/Zonula occludens-1 (PDZ)-domain is involved in recognition of and interaction between various proteins, by which the localization and the regulation are mediated. Multi-PDZ domain protein 1 (MUPP1) contains 13 PDZ domains. MUPP1 serves a scaffolding function for structure proteins and signaling proteins, but the mechanism how MUPP1 is stabilized and signalized has not yet been elucidated. We used the yeast two-hybrid system to identify proteins that interact with PDZ domains of MUPP1. We found an interaction between MUPP1 and Parkin. Parkin is an E3 ubiquitin ligase. Loss-of-function mutations of Parkin gene are known to cause an autosomal recessive juvenile parkinsonism. Parkin bound to the $12^{th}$ PDZ domain, but not to other PDZ domains of MUPP1. The C-terminal end of Parkin has a type II PDZ-association motif, which was essential for the interaction with MUPP1 in the yeast two-hybrid assay. When co-expressed in HEK-293T cells, Parkin co-localized with MUPP1. When co-expressed with ubiquitin in HEK-293T cells, MUPP1 has been strongly ubiquitinated by Parkin. These findings collectively suggest that MUPP1 is a novel substrate of Parkin and its function or stability could be modulated by Parkin-mediated ubiquitination.

• Journal of Life Science
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• v.26 no.9
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• pp.991-998
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• 2016

NF-κB acts as a critical transcription factor for the survival of cells via the induction of antiapoptotic genes. Constitutive activation of NF-κB in many types of solid tumors suggests that the inhibition of NF-κB might prevent or inhibit tumorigenesis. Although a number of studies demonstrated that Hsp70 regulated NF-κB activity, the exact mechanism is not clear. This study investigated the functional relationship of Hsp70 and IKKγ in the regulation of NF-κB activation using expression plasmids of components of the IKK complex. Wild-type and deletion mutants of IKKγ were expressed together with Hsp70, and the combined regulatory effect of Hsp70 and IKKγ on NF-κB activation was assayed. Hsp70 suppressed the activation of NF-κB in a reporter plasmid assay. Hsp70 also suppressed the phosphorylation and degradation of IκBα. The suppressive effect of Hsp70 on NF-κB activation was synergistically elevated by IKKγ. The N-terminal IKKβ binding site, C-terminal leucine zipper, and zinc finger domains of IKKγ were not necessary for the suppressive effect. Furthermore, Hsp70 and IKKγ synergistically suppressed the induction of COX-2 expression by lipopolysaccharides in RAW264.7 cells. These results suggest that overexpression of Hsp70 and IKKγ may be a strategic method for inhibition of NF-κB and related diseases.

• Journal of Life Science
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• v.26 no.9
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• pp.1056-1062
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• 2016

Lung cancer is currently the most common malignant disease and the leading cause of mortality in the world and non-small cell lung cancer (NSCLC) accounts for 75-80% of lung cancer cases. miR-155 gene was found to be over expressed in several solid tumors, such as thyroid carcinoma, breast cancer, colon cancer, cervical cancer, pancreatic ductal adenocarcinoma (PDAC) and lung cancer. The aims of this study were to define the expression of miR-155 in lung cancer and its associated clinic-pathologic characteristics. Total RNA was purified from formalin-fixed, paraffin-embedded NSCLC tissues and benign lung tissues. Expression of miR-155 in human lung cancer tissues were evaluated as mean fold changes of miR-155 in cancer tissues compared to benign lung tissues by quantitative real-time reverse transcriptase polymerase chain reaction (real-time qRT-PCR) and associations of miR-155 expression with clinic-pathologic findings of cancer. Compared with the benign control group, miR-155 expression was significantly overexpressed in NSCLCs (p=<0.001). miR-155 was more overexpressed in squamous cell carcinoma than in adenocarcinoma. Poorly differentiated tumors showed significantly overexpression of miR-155 than well-differentiated tumors (p=<0.001). Overexpression of miR-155 was significantly associated with lymph node metastasis (p=<0.05). In survival analysis for all NSCLC patients, high miR-155 expression was significantly correlated with worse overall survival (p=<0.05). These results suggested that miR-155 might play an important role in lung cancer progression and metastasis.

• Korean Journal of Plant Resources
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• v.29 no.1
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• pp.1-10
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• 2016

Obesity is a pro-inflammatory state that contributes to the development of metabolic disorders such as hyperlipidemia, insulin resistance, type 2 diabetes, non-alcoholic fatty liver, and cardiovascular disease. In this study, we evaluated the inhibition of adipogenesis in 3T3-L1 cells and in high-fat diet (HFD)-induced obese mice by Peucedanum japonicum Thunberg L. water extract (PJT). Lipid accumulation measurement indicates that PJT markedly inhibited adipogenesis in a dose-dependent manner. RT-PCR results demonstrated that the mRNA expression of adipogenic transcription factors such as peroxisome proliferator-activated receptor-γ (PPARγ) and CCAAT/enhancer binding protein-α (C/EBPα) in 3T3-L1 cells were significantly down-regulated by PJT treatment. Oral administration of PJT (100, 300, and 500 ㎎/㎏, b.w/daily for 4 weeks) was conducted in high-fat diet induced obese mice and C57BL/6 mice. The PJT-administered group of HFD-induced mice had a lower body weight gain, along with decreased serum levels of glucose, triglycerides, and total cholesterol compared with the control mice, however, the HDL-cholesterol/total cholesterol ratio was increased. Furthermore, the elevated mRNA expression levels of adipogenesis related genes in the white adipose tissue of obese mice were significantly suppressed by PJT. These results indicate that PJT exhibits anti-obesity effects in obese mice by decreasing in serum lipid levels and lipogenesis related gene.