Korea Research Institute of Bioscience and Biotechnology
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Anti-proliferative effect of Allium senescens L. extract in human T-cell acute lymphocytic leukemia cells
No full textAllium species are well known plants distributed throughout the world, and they contain various bioactive components with different biological activities including anti-cancer effects. In this study, we investigated the inhibitory effect of Allium senescens L. (A.S.) extract on cell survival and IL-2-mediated inflammation in human T cell acute lymphocytic leukemia (T-ALL) Jurkat cells. Our results showed that A.S. extract induced caspase-dependent apoptosis of Jurkat cells with no significant cytotoxicity in the normal peripheral blood mononuclear cells. A.S. extract induced ROS generation through the activation of MAPK p38 phosphorylation. It also inhibited IL-2 mRNA expression and NF-κB signaling mediated by phorbol 12-myristate 13-acetate, and phytohemagglutinin. Combined treatment with A.S. extract and axitinib/dovitinib exerted enhanced inhibitory effects on T-ALL cell growth and IL-2 production. These results provide novel information on the potential use of A.S. extract as a therapeutic herbal agent for the treatment and prevention of T-ALL.
Overexpression of the golden SNP-carrying Orange gene enhances carotenoid accumulation and heat stress tolerance in sweetpotato plants
No full textCarotenoids function as photosynthetic accessory pigments, antioxidants, and vitamin A precursors. We recently showed that transgenic sweetpotato calli overexpressing the mutant sweetpotato (Ipomoea batatas [L.] Lam) Orange gene (IbOr-R96H), which carries a single nucleotide polymorphism responsible for Arg to His substitution at amino acid position 96, exhibited dramatically higher carotenoid content and abiotic stress tolerance than calli overexpressing the wild-type IbOr gene (IbOr-WT). In this study, we generated transgenic sweetpotato plants overexpressing IbOr-R96H under the control of the cauliflower mosaic virus (CaMV) 35S promoter via Agrobacterium-mediated transformation. The total carotenoid contents of IbOr-R96H storage roots (light-orange flesh) and IbOr-WT storage roots (light-yellow flesh) were 5.4？19.6 and 3.2-fold higher, respectively, than those of non-transgenic (NT) storage roots (white flesh). The β-carotene content of IbOr-R96H storage roots was up to 186.2-fold higher than that of NT storage roots. In addition, IbOr-R96H plants showed greater tolerance to heat stress (47 °C) than NT and IbOr-WT plants, possibly because of higher DPPH radical scavenging activity and ABA contents. These results indicate that IbOr-R96H is a promising strategy for developing new sweetpotato cultivars with improved carotenoid contents and heat stress tolerance.
Biosensor-based directed evolution of methanol dehydrogenase from Lysinibacillus xylanilyticus
No full textMethanol dehydrogenase (Mdh), is a crucial enzyme for utilizing methane and methanol as carbon and energy sources in methylotrophy and synthetic methylotrophy. Engineering of Mdh, especially NAD-dependent Mdh, has thus been actively investigated to enhance methanol conversion. However, its poor catalytic activity and low methanol affinity limit its wider application. In this study, we applied a transcriptional factor-based biosensor for the direct evolution of Mdh from Lysinibacillus xylanilyticus (Lxmdh), which has a relatively high turnover rate and low KM value compared to other wild-type NAD-dependent Mdhs. A random mutant library of Lxmdh was constructed in Escherichia coli and was screened using formaldehyde-detectable biosensors by incubation with low methanol concentrations. Positive clones showing higher fluorescence were selected by fluorescence-activated cell sorting (FACS) system, and their catalytic activities toward methanol were evaluated. The successfully isolated mutants E396V, K318N, and K46E showed high activity, particularly at very low methanol concentrations. In kinetic analysis, mutant E396V, K318N, and K46E had superior methanol conversion efficiency, with 79-, 23-, and 3-fold improvements compared to the wild-type, respectively. These mutant enzymes could thus be useful for engineering synthetic methylotrophy and for enhancing methanol conversion to various useful products.
Overexpression of the sweetpotato peroxidase gene swpa4 enhances tolerance to methyl viologen-mediated oxidative stress and dehydration in Arabidopsis thaliana
No full textWe previously reported that transgenic Arabidopsis thaliana plants overexpressing the sweet potato peroxidase gene swpa4 under the control of the cauliflower mosaic virus (CaMV) 35 s promoter showed increased levels of reactive oxygen species (ROS) and nitric oxide (NO), and higher expression of ROS and NO related genes than control plants. Here, we investigated the effect of swpa4 overexpression on the abiotic and biotic stress tolerance levels of Arabidopsis plants. Methyl viologen (MV) treatment-induced oxidative stress caused visible damage to the seedlings and rosette leaves of all Arabidopsis plants, although the symptoms were more severe in control plants than in transgenic lines. Additionally, survival rates and ion leakage showed a slight decline in transgenic lines but a more severe decline in control plants after MV treatment. Transgenic plants also showed enhanced tolerance to drought stress. Dehydration treatment, followed by rehydration, resulted in a greater change in the relative water content and lipid peroxidation of control plants than in that of transgenic lines. However, transgenic plants did not show enhanced resistance to biotic stresses such as bacterial and fungal pathogens. These results indicate that transgenic Arabidopsis plants can efficiently regulate defense levels during oxidative stress via the overexpression of swpa4.
Dutomycin induces autophagy and apoptosis by targeting the serine protease inhibitor SERPINB6
No full textAutophagy plays an important role in maintaining tumor cell progression and survival in response to metabolic stress. Thus, the regulation of autophagy can be used as a strategy for anticancer therapy. Here, we report dutomycin (DTM) as a novel autophagy enhancer that eventually induces apoptosis due to excessive autophagy. Also, human serine protease inhibitor B6 (SERPINB6) was identified as a target protein of DTM, and its novel function which is involved in autophagy was studied for the first time. We show that DTM directly binds SERPINB6 and then activates intracellular serine proteases, resulting in autophagy induction. Inhibitory effects of DTM on the function of SERPINB6 were confirmed through enzyme- and cell-based approaches, and SERPINB6 was validated as a target protein using siRNA-mediated knockdown and an overexpression test. In a zebrafish xenograft model, DTM showed a significant decrease in tumor area. Furthermore, the present findings will be expected to contribute to the expansion of novel basic knowledge about the correlation of cancer and autophagy by promoting active further research on SERPINB6, which was not previously considered the subject of cancer biology.
Establishment of iPSC (KRIBBi001-A) from CD34+ group O D-negative bone marrow blood
No full textHuman induced pluripotent stem cells with indefinite propagation in vitro provide a potential donor source of cells including erythroid cells for human therapy. Since group O D-negative (RhD？) blood cells are considered as universal donors for transfusion, it is compelling to derive iPSC line from group O/RhD？ sample as a new cellular source to generate universal RBCs. The resulting iPSC line derived from group O/RhD？ somatic source showed typical features of pluripotent stem cells and could provide an unprecedented cellular tool to develop universal therapeutics for blood transfusion.
TNFα enhances trovafloxacin-induced in vitro hepatotoxicity by inhibiting protective autophagy
No full textTrovafloxacin (TVX) is associated with idiosyncratic drug-induced liver injury (iDILI) and inflammation-mediated hepatotoxicity. However, the inflammatory stress-regulated mechanisms in iDILI remain unclear. Herein, we elucidated the novel role of tumor-necrosis factor alpha (TNFα), an inflammatory stress factor, in TVX-induced in vitro hepatotoxicity and synergistic toxicity. TVX specifically induced synergistic toxicity in HepG2 cells with TNFα, which inhibits autophagy. TVX-treated HepG2 cells induced protective autophagy by inhibiting the expression of mTOR signaling proteins, while ATG5 knockdown in HepG2 cells, responsible for the impairment of autophagy, enhanced TVX-induced toxicity due to the increase in cytochrome C release and JNK pathway activation. Interestingly, the expression of mTOR signal proteins, which were suppressed by TVX, disrupted the negative feedback of the PI3K/AKT pathway and TNFα rebounded p70S6K phosphorylation. Co-treatment with TVX and TNFα inhibited protective autophagy by maintaining p70S6K activity, which enhanced TVX-induced cytotoxicity. Phosphorylation of p70S6K was inhibited by siRNA knockdown and rapamycin to restore TNFα-inhibited autophagy, which prevented the synergistic effect on TVX-induced cytotoxicity. These results indicate that TVX activates protective autophagy in HepG2 cells exposed to toxicity and an imbalance in negative feedback regulation of autophagy by TNFα synergistically enhanced the toxicity. The finding from this study may contribute to a better understanding of the mechanisms underlying iDILI associated with inflammatory stress.
Influenza chimeric protein (3M2e-3HA2-NP) adjuvanted with PGA/alum conferred cross-protection against heterologous influenza A viruses
No full textVaccination is the most effective way to prevent influenza virus infections. However, conventional vaccines based on hemagglutinin (HA) have to be annually updated because the HA of influenza viruses constantly mutates. In this study, we produced a 3M2e？3HA2？NP chimeric protein as a vaccine antigen candidate using an Escherichia coli expression system. The vaccination of chimeric protein (15 μg) conferred complete protection against A/Puerto Rico/8/1934 (H1N1; PR8) in mice. It strongly induced influenza virus-specific antibody responses, cytotoxic T lymphocyte activity, and antibody-dependent cellular cytotoxicity. To spare the dose and enhance the cross-reactivity of the chimeric, we used a complex of poly-γ-glutamic acid and alum (PGA/alum) as an adjuvant. PGA/alumadjuvanted, low-dose chimeric protein (1 or 5 μg) exhibited higher cross-protective effects against influenza A viruses (PR8, CA04, and H3N2) compared with those of chimeric alone or alumadjuvanted proteins in vaccinated mice. Moreover, the depletion of CD4+ T, CD8+ T, and NK cells reduced the survival rate and efficacy of the PGA/alum-adjuvanted chimeric protein. Collectively, the vaccination of PGA/alum-adjuvanted chimeric protein induced strong protection efficacy against homologous and heterologous influenza viruses in mice, which suggests that it may be a promising universal influenza vaccine candidate.
Effects of post-fire vegetation recovery on soil erosion in vulnerable montane regions in a monsoon climate: a decade of monitoring
No full textIn areas with frequent fires and erosive soils under monsoon climate, we aimed to determine the long-term dynamics of soil loss during vegetation recovery and to examine elapsed time for soil stabilization. Runoff plots were installed in Pinus densiflora forests affected by stand-replacing fires on the East Coast of South Korea, which occurred in spring 2000. Runoff plots measured runoff and sediment yields for 11 years (2003？2013) during which time, vegetation cover of low-, intermediate-, and high-vegetation cover plots increased from 21 to 44%, 49% to 69%, and 87% to 95%, respectively. Vegetation was effective in preventing runoff and sediment yield. Nevertheless, to stabilize to below baseline load, it took 7 years at low and intermediate cover plots and only 3 years at high cover plots. For 7 years, 7.06 (5.2-fold) and 4.29 (3.1-fold) kg m？2 of soil were lost at the low and intermediate cover plots, respectively, compared to the high cover plots (1.37 kg m？2). Sediment loss fluctuated more than runoff following extreme rainfall events. We suggest that for the slow recovery area (？70% cover).
Industrialization of a novel functional cosmetic material based on marine microalge oil
No full text해양미세조류 유래 오일 기반 기능성 화장품 소재 산업화MBC005201