Korea Research Institute of Bioscience and Biotechnology
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Analysis and stability test of the extracts from ulmus parvifolia leaves
No full textA simple and reliable HPLC method was developed to determine pharmacologically standard marker compounds of Ulmus parvifolia leaves. Standard markers were characterized with neochlorogenic acid (trans-5-O-caffeoylquinic acid, 5-CQA) and chlorogenic acid (trans-3-O-caffeoylquinic acid, 3-CQA) using NMR and UPLC-QTof-MS analysis. A method for qualitative/quantitative analysis of the leaves extracts were evaluated including two compounds by using HPLC. The stability test of 30% ethanolic extracts of the leaves sample and standard markers have been evaluated for six months. However, no significant changes in the content of the marker compounds of each extract was observed during the time of investigation.
Comparative transcriptome profiling of tuberous roots of two sweetpotato lines with contrasting low temperature tolerance during storage
No full textSweetpotato (Ipomoea batatas [L.] Lam) is considered an economically important crop worldwide and is used as a source of food, feed, and biomaterials. However, its origin in tropical regions makes it vulnerable to chilling injury during postharvest storage at low temperature. To gain further insight into the molecular mechanism of chilling response, we performed comparative transcriptome analysis of two sweetpotato lines, Xushu 15-1 and Xushu 15-4, with high and low cold storage ability, respectively. Tuberous roots of these lines were stored at 4？°C for 0, 2, and 6？weeks. RNA-Seq data of both lines were de novo assembled, producing 27,636 unigenes with a N50 value of 1204？bp. A total of 525 differentially expressed genes (DEGs) were identified and categorized into six clusters. Genes with higher expression in Xushu 15-1 than in Xushu 15-4 significantly increased in number over time during low temperature storage. Functional annotation of DEGs using KEGG enrichment analysis showed that these DEGs were involved in carbohydrate metabolism, ribosome, protein processing in endoplasmic reticulum, plant-pathogen interaction, and plant hormone signal transduction. Several key candidate genes involved in KEGG pathways were selected and discussed further. The results of this study enhance our understanding of the complex mechanisms involved in low temperature tolerance in sweetpotato during storage and provide a set of candidate genes for the development of new varieties with improved cold storage ability.
Production of adipic acid by short- and long-chain fatty acid acyl-CoA oxidase engineered in yeast Candida tropicalis
No full textIn this study, to produce adipic acid, mutant strains of Candida tropicalis KCTC 7212 deficient of AOX genes encoding acyl-CoA oxidases which are important in the β-oxidation pathway were constructed. Production of adipic acid in the mutants from the most favorable substrate C12 methyl laurate was significantly increased. The highest level of production of adipic acid was obtained in the C. tropicalis ΔAOX4::AOX5 mutant of 339.8 mg L-1 which was about 5.4-fold higher level compared to the parent strain. The C. tropicalis ΔAOX4::AOX5 mutant was subjected to fed-batch fermentation at optimized conditions of agitation rate of 1000 rpm, pH 5.0 and methyl laurate of 3% (w/v), giving the maximum level of adipic acid of 12.1 g L-1 and production rate of 0.1 g L-1 h-1.
Quercetin attenuates decrease of thioredoxin expression following focal cerebral ischemia and glutamate-induced neuronal cell damage
No full textQuercetin is a bioactive flavonoid which abundantly exists in vegetables and fruits. Quercetin exerts a neuroprotective effect against cerebral ischemia. Thioredoxin acts as antioxidant by regulating redox signaling. This study investigated whether quercetin regulates thioredoxin expression in focal cerebral ischemia and glutamate-induced neuronal cell death. Male Sprague Dawley rats (210-230？g) were intraperitoneally injected with vehicle or quercetin (10？mg/kg) 1？h prior to middle cerebral artery occlusion (MCAO). Cerebral cortex was collected 24？h after MCAO. MCAO led to neurological movement deficits, brain edema, and serious histopathological damages in cerebral cortex, and quercetin alleviated these damages following MCAO. We observed the change of thioredoxin expression in MCAO animals with quercetin using proteomic approach, reverse-transcription PCR, and Western blot analyses. Thioredoxin expression decreased in vehicle-treated MCAO animals, while quercetin attenuated this decrease. Moreover, quercetin treatment alleviated the decrease in the number of thioredoxin-positive cells in cerebral cortex of MCAO animals. Furthermore, immunoprecipitation analysis demonstrated that interaction of apoptosis signal-regulating kinase 1 (ASK1) and thioredoxin was decreased in MCAO animals with vehicle, while quercetin prevented MCAO-induced decrease in these binding. In addition, quercetin also alleviated the reduction of cell viability and the decrease in thioredoxin expression in glutamate-treated hippocampal cell line and primary cultures of cortical neurons. However in thioredoxin-silenced cortical neuron, anti-apoptotic effect of quercetin was decreased. Thus, changes of thioredoxin expression by quercetin may contribute to the neuroprotective effect of quercetin in focal cerebral ischemia. Our findings suggest that quercetin mediates its neuroprotective function by regulation of thioredoxin expression and maintenance of interaction between ASK1 and thioredoxin.
Regulation of PTP1B activation through disruption of redox-complex formation
No full textWe have identified a molecular interaction between the reversibly oxidized form of protein tyrosine phosphatase 1B (PTP1B) and 14-3-3ζ that regulates PTP1B activity. Destabilizing the transient interaction between 14-3-3ζ and PTP1B prevented PTP1B inactivation by reactive oxygen species and decreased epidermal growth factor receptor phosphorylation. Our data suggest that destabilizing the interaction between 14-3-3ζ and the reversibly oxidized and inactive form of PTP1B may establish a path to PTP1B activation in cells.
Golgi-localized cyclophilin 21 proteins negatively regulate ABA signalling via the peptidyl prolyl isomerase activity during early seedling development
No full textPlant possesses particular Golgi-resident cyclophilin 21 proteins (CYP21s) and the catalytic isomerase activities have a negative effect on ABA signalling gene expression during early seedling development. Cyclophilins (CYPs) are essential for diverse cellular process, as these catalyse a rate-limiting step in protein folding. Although Golgi proteomics in Arabidopsis thaliana suggests the existence of several CYPs in the Golgi apparatus, only one putative Golgi-resident CYP protein has been reported in rice (Oryza sativa L.; OsCYP21-4). Here, we identified the Golgi-resident CYP21 family genes and analysed their molecular characteristics in Arabidopsis and rice. The CYP family genes (CYP21-1, CYP21-2, CYP21-3, and CYP21-4) are plant-specific, and their appearance and copy numbers differ among plant species. CYP21-1 and CYP21-4 are common to all angiosperms, whereas CYP21-2 and CYP21-3 evolved in the Malvidae subclass. Furthermore, all CYP21 proteins localize to cis-Golgi, trans-Golgi or both cis- and trans-Golgi membranes in plant cells. Additionally, based on the structure, enzymatic function, and topological orientation in Golgi membranes, CYP21 proteins are divided into two groups. Genetic analysis revealed that Group I proteins (CYP21-1 and CYP21-2) exhibit peptidyl prolyl cis-trans isomerase (PPIase) activity and regulate seed germination and seedling growth and development by affecting the expression levels of abscisic acid signalling genes. Thus, we identified the Golgi-resident CYPs and demonstrated that their PPIase activities are required for early seedling growth and development in higher plants.
Changes in the expression of mitochondrial morphology-related genes during the differentiation of murine embryonic stem cells
No full textDuring embryonic development, cells undergo changes in gene expression, signaling pathway activation/inactivation, metabolism, and intracellular organelle structures, which are mediated by mitochondria. Mitochondria continuously switch their morphology between elongated tubular and fragmented globular via mitochondrial fusion and fission. Mitochondrial fusion is mediated by proteins encoded by Mfn1, Mfn2, and Opa1, whereas mitochondrial fission is mediated by proteins encoded by Fis1 and Dnm1L. Here, we investigated the expression patterns of mitochondria-related genes during the differentiation of mouse embryonic stem cells (ESCs). Pluripotent ESCs maintain stemness in the presence of leukemia inhibitory factor (LIF) via the JAK-STAT3 pathway but lose pluripotency and differentiate in response to the withdrawal of LIF. We analyzed the expression levels of mitochondrial fusion- and fission-related genes during the differentiation of ESCs. We hypothesized that mitochondrial fusion genes would be overexpressed while the fission genes would be downregulated during the differentiation of ESCs. Though the mitochondria exhibited an elongated morphology in ESCs differentiating in response to LIF withdrawal, only the expression of Mfn2 was increased and that of Dnm1L was decreased as expected, the other exceptions being Mfn1, Opa1, and Fis1. Next, by comparing gene expression and mitochondrial morphology, we proposed an index that could precisely represent mitochondrial changes during the differentiation of pluripotent stem cells by analyzing the expression ratios of three fusion- and two fission-related genes. Surprisingly, increased Mfn2/Dnm1L ratio was correlated with elongation of mitochondria during the differentiation of ESCs. Moreover, application of this index to other specialized cell types revealed that neural stems cells (NSCs) and mouse embryonic fibroblasts (MEFs) showed increased Mfn2/Dnm1L ratio compared to ESCs. Thus, we suggest that the Mfn2/Dnm1L ratio could reflect changes in mitochondrial morphology according to the extent of differentiation.
Paenibacillus oralis sp. nov., isolated from human subgingival dental plaque of gingivitis lesion
No full textA Gram-stain-negative, facultative anaerobic, spore-forming, motile, and rod-shaped bacterium, strain ChDC PVNT-B20T, was isolated from the human subgingival dental plaque of a gingivitis lesion. Phylogenetic analysis based on the 16S ribosomal RNA gene (16S rDNA) showed that the strain belonged to the genus Paenibacillus. BLAST analysis of 16S rDNA sequence of the strain displayed high identity to those of Paenibacillus faecis DSM 23593T (97.7% similarity) and Paenibacillus macerans ATCC 8244T (97.6% similarity). Draft genome of strain ChDC PVNT-B20T was composed of 8,112,407 bp. The DNA G+C content of the strain was 51.3 mol%. Average nucleotide identity values between strain ChDC PVNT-B20T and P. faecis DSM 23593T or P. macerans ATCC 8244T were 75.71% and 91.5%, respectively. Genome-to-genome distance values between strain ChDC PVNT-B20T and P. faecis DSM 23593T or P. macerans ATCC 8244T were 21.6% (19.3-24.0%) and 44.9% (42.3-47.4%), respectively. Major cellular fatty acids of strain ChDC PVNT-B20T were anteiso-C15:0 (43.4%), C16:0 (16.6%), iso-C16:0 (14.4%), and anteiso-C17:0 (12.4%). The sole respiratory quinone of the strain was menaqinone-7. Major polar lipids of the strain were phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), and one unidentified glycolipid (GL). Minor polar lipids were one unidentified aminolipid (AL), one unidentified phospholipid (PL), and three unidentified lipids (L1-L3). Based on these results, strain ChDC PVNT-B20T is considered to represent a novel species of the genus Paenibacillus, for which the name Paenibacillus oralis sp. nov. is proposed. Type strain is ChDC PVNT-B20T (=？KCOM 3021T？=？JCM 33462 T).
Spiraea prunifolia var. simpliciflora attenuates oxidative stress and inflammatory responses in a murine model of lipopolysaccharide-iInduced acute lung injury and TNF-α-stimulated NCI-H292 cells
No full textSpiraea prunifolia var. simpliciflora (SP) is traditionally used as an herbal remedy to treat fever, malaria, and emesis. This study aimed to evaluate the anti-oxidative and anti-inflammatory properties of the methanol extract of SP leaves in tumor necrosis factor (TNF)-α-stimulated NCI-H292 cells and in a lipopolysaccharide (LPS)-induced acute lung injury (ALI) mouse model. SP decreased the number of inflammatory cells and the levels of TNF-α, interleukin (IL)-1β, and IL-6 in the bronchoalveolar lavage fluid, and inflammatory cell infiltration in the lung tissues of SP-treated mice. In addition, SP significantly suppressed the mRNA and protein levels of TNF-α, IL-1β, and IL-6 in TNF-α-stimulated NCI-H292 cells. SP significantly suppressed the phosphorylation of the mitogen-activated protein kinases (MAPKs) and p65-nuclear factor-kappa B (NF-κB) in LPS-induced ALI mice and TNF-α-stimulated NCI-H292 cells. SP treatment enhanced the nuclear translocation of nuclear factor erythroid 2-related factor (Nrf2) with upregulated antioxidant enzymes and suppressed reactive oxygen species (ROS)-mediated oxidative stress in the lung tissues of LPS-induced ALI model and TNF-α-stimulated NCI-H292 cells. Collectively, SP effectively inhibited airway inflammation and ROS-mediated oxidative stress, which was closely related to its ability to induce activation of Nrf2 and inhibit the phosphorylation of MAPKs and NF-κB. These findings suggest that SP has therapeutic potential for the treatment of ALI.
A TMT-based quantitative proteome analysis to elucidate the TSWV induced signaling cascade in susceptible and resistant cultivars of Solanum lycopersicum
No full textTomato spotted wilt virus (TSWV), transmitted by small insects known as thrips, is one of the major threats to tomato productivity across the globe. In addition to tomato, this virus infects more than 1000 other plants belonging to 85 families and is a cause of serious concern. Very little, however, is known about the molecular mechanism of TSWV induced signaling in plants. Here, we used a tandem mass tags (TMT)-based quantitative proteome approach to investigate the protein profiles of tomato leaves of two cultivars (cv 2621 and 2689; susceptible and resistant to TSWV infection, respectively) following TSWV inoculation. This approach resulted in the identification of 5112 proteins of which 1022 showed significant changes in response to TSWV. While the proteome of resistant cultivar majorly remains unaltered, the proteome of susceptible cultivar showed distinct differences following TSWV inoculation. TSWV modulated proteins in tomato included those with functions previously implicated in plant defense including secondary metabolism, reactive oxygen species (ROS) detoxification, mitogen-activated protein (MAP) kinase signaling, calcium signaling and jasmonate biosynthesis, among others. Taken together, results reported here provide new insights into the TSWV induced signaling in tomato leaves and may be useful in the future to manage this deadly disease of plants.