• Molecules and Cells
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• v.40 no.10
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• pp.773-786
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• 2017

The loss of green coloration via chlorophyll (Chl) degradation typically occurs during leaf senescence. To date, many Chl catabolic enzymes have been identified and shown to interact with light harvesting complex II to form a Chl degradation complex in senescing chloroplasts; this complex might metabolically channel phototoxic Chl catabolic intermediates to prevent oxidative damage to cells. The Chl catabolic enzyme 7-hydroxymethyl Chl a reductase (HCAR) converts 7-hydroxymethyl Chl a (7-HMC a) to Chl a. The rice (Oryza sativa) genome contains a single HCAR homolog (OsHCAR), but its exact role remains unknown. Here, we show that an oshcar knockout mutant exhibits persistent green leaves during both dark-induced and natural senescence, and accumulates 7-HMC a and pheophorbide a (Pheo a) in green leaf blades. Interestingly, both rice and Arabidopsis hcar mutants exhibit severe cell death at the vegetative stage; this cell death largely occurs in a light intensity-dependent manner. In addition, 7-HMC a treatment led to the generation of singlet oxygen ($^1O_2$) in Arabidopsis and rice protoplasts in the light. Under herbicide-induced oxidative stress conditions, leaf necrosis was more severe in hcar plants than in wild type, and HCAR-overexpressing plants were more tolerant to reactive oxygen species than wild type. Therefore, in addition to functioning in the conversion of 7-HMC a to Chl a in senescent leaves, HCAR may play a critical role in protecting plants from high light-induced damage by preventing the accumulation of 7-HMC a and Pheo a in developing and mature leaves at the vegetative stage.

• Natural Product Sciences
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• v.22 no.3
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• pp.201-208
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• 2016

Here in this study, we investigated the lifespan-extending effect and underlying mechanism of methanolic extract of Moringa olelifa leaves (MML) using Caenorhabditis elegans (C. elegans) model system. To define the longevity properties of MML we conducted lifespan assay and MML showed significant increase in lifespan under normal culture condition. In addition, MML elevated stress tolerance of C. elegans to endure against thermal, oxidative and osmotic stress conditions. Our data also revealed that increased activities of antioxidant enzymes and expressions of stress resistance proteins were attributed to MML-mediated enhanced stress resistance. We further investigated the involvement of MML on the aging-related factors such as growth, food intake, fertility, and motility. Interestingly, MML significantly reduced growth and egg-laying, suggesting these factors were closely linked with MML-mediated longevity. We also observed the movement of aged worms to estimate the effects of MML on the health span. Herein, MML efficiently elevated motility of aged worms, indicating MML may affect health span as well as lifespan. Our genetic analysis using knockout mutants showed that lifespan-extension activity of MML was interconnected with several genes such as skn-1, sir-2.1, daf-2, age-1 and daf-16. Based on these results, we could conclude that MML prolongs the lifespan of worms via activation of SKN-1 and SIR-2.1 and inhibition of insulin/IGF pathway, followed by DAF-16 activation.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1176-1182
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• 2018

Objective: Progesterone receptor (PGR) and AT-rich interactive domain 1A (ARID1A) have important roles in the establishment and maintenance of pregnancy in the uterus. In present studies, we examined the expression of mitochondrial tumor suppressor 1 (MTUS1) in the murine uterus during early pregnancy as well as in response to ovarian steroid hormone treatment. Methods: We performed quantitative reverse transcription polymerase chain reaction and immunohistochemistry analysis to investigate the regulation of MTUS1 by ARID1A and determined expression patterns of MTUS1 in the uterus during early pregnancy. Results: The expression of MTUS1 was detected on day 0.5 of gestation (GD 0.5) and then gradually increased until GD 3.5 in the luminal and glandular epithelium. However, the expression of MTUS1 was significantly reduced in the uterine epithelial cells of $Pgr^{cre/+}Arid1a^{f/f}$ and Pgr knockout (PRKO) mice at GD 3.5. Furthermore, MTUS1 expression was remarkably induced after P4 treatment in the luminal and glandular epithelium of the wild-type mice. However, the induction of MTUS1 expression was not detected in uteri of $Pgr^{cre/+}Arid1a^{f/f}$ or PRKO mice treated with P4. Conclusion: These results suggest that MTUS1 is a novel target gene by ARID1A and PGR in the uterine epithelial cells.

• Journal of Microbiology and Biotechnology
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• v.20 no.12
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• pp.1630-1636
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• 2010

During the fermentation process of Saccharomyces cerevisiae, yeast cells must rapidly respond to a wide variety of external stresses in order to survive the constantly changing environment, including ethanol stress. The accumulation of ethanol can severely inhibit cell growth activity and productivity. Thus, the response to changing ethanol concentrations is one of the most important stress reactions in S. cerevisiae and worthy of thorough investigation. Therefore, this study examined the relationship between ethanol tolerance in S. cerevisiae and a unique protein called alcohol sensitive RING/PHD finger 1 protein (Asr1p). A real-time PCR showed that upon exposure to 8% ethanol, the expression of Asr1 was continuously enhanced, reaching a peak 2 h after stimulation. This result was confirmed by monitoring the fluorescence levels using a strain with a green fluorescent protein tagged to the C-terminal of Asr1p. The fluorescent microscopy also revealed a change in the subcellular localization before and after stimulation. Furthermore, the disruption of the Asr1 gene resulted in hypersensitivity on the medium containing ethanol, when compared with the wild-type strain. Thus, when taken together, the present results suggest that Asr1 is involved in the response to ethanol stress in the yeast S. cerevisiae.

• Molecules and Cells
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• v.38 no.12
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• pp.1111-1117
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• 2015

Toll-like receptor 5 (TLR5) is a specific receptor for microbial flagellin and is one of the most well-known receptors in the TLR family. We reported previously that TLR5 signaling is well maintained during aging and that caveolin-1 may be involved in TLR5 signaling in aged macrophages through direct interactions. Therefore, it is important to clarify whether caveolin-1/TLR5 interactions affect TLR5 expression during aging. To assess the effect of caveolin-1 on TLR5, we analyzed TLR5 expression in senescent fibroblasts and aged tissues expressing high levels of caveolin-1. As expected, TLR5 mRNA and protein expression was well maintained in senescent fibroblasts and aged tissues, whereas TLR4 mRNA and protein were diminished in those cells and tissues. To determine the mechanism of caveolin-1-dependent TLR5 expression, we examined TLR5 expression in caveolin-1 deficient mice. Interestingly, TLR5 mRNA and protein levels were decreased dramatically in tissues from caveolin-1 knockout mice. Moreover, overexpressed caveolin-1 in vitro enhanced TLR5 mRNA through the MAPK pathway and prolonged TLR5 protein half-life through direct interaction. These results suggest that caveolin-1 may play a crucial role in maintaining of TLR5 by regulating transcription systems and increasing protein half-life.

• Molecules and Cells
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• v.40 no.8
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• pp.577-586
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• 2017

Phytocystatins (PhyCYSs) are plant-specific proteinaceous inhibitors that are implicated in protein turnover and stress responses. Here, we characterized a PhyCYS from Arabidopsis thaliana, which was designated AtCYS5. RT-qPCR analysis showed that the expression of AtCYS5 in germinating seeds was induced by heat stress (HS) and exogenous abscisic acid (ABA) treatment. Analysis of the expression of the ${\beta}-glucuronidase$ reporter gene under the control of the AtCYS5 promoter showed that AtCYS5 expression during seed germination was induced by HS and ABA. Constitutive overexpression of AtCYS5 driven by the cauliflower mosaic virus 35S promoter led to enhanced HS tolerance in transgenic Arabidopsis, which was characterized by higher fresh weight and root length compared to wild-type (WT) and knockout (cys5) plants grown under HS conditions. The HS tolerance of AtCYS5-overexpressing transgenic plants was associated with increased insensitivity to exogenous ABA during both seed germination and post-germination compared to WT and cys5. Although no HS elements were identified in the 5'-flanking region of AtCYS5, canonical ABA-responsive elements (ABREs) were detected. AtCYS5 was upregulated in ABAtreated protoplasts transiently co-expressing this gene and genes encoding bZIP ABRE-binding factors (ABFs and AREB3). In the absence of ABA, ABF1 and ABF3 directly bound to the ABREs in the AtCYS5 promoter, which activated the transcription of this gene in the presence of ABA. These results suggest that an ABA-dependent pathway plays a positive role in the HS-responsive expression of AtCYS5 during seed germination and post-germination growth.

• Molecules and Cells
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• v.41 no.2
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• pp.140-149
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• 2018

The $TIS21^{/BTG2/PC3}$ gene belongs to the antiproliferative gene (APRO) family and exhibits tumor suppressive activity. However, here we report that TIS21 controls lipid metabolism, rather than cell proliferation, under fasting condition. Using microarray analysis, whole gene expression changes were investigated in liver of TIS21 knockout (TIS21-KO) mice after 20 h fasting and compared with wild type (WT). Peroxisome proliferator-activated receptor alpha ($PPAR{\alpha}$) target gene expression was almost absent in contrast to increased lipid synthesis in the TIS21-KO mice compared to WT mice. Immunohistochemistry with hematoxylin and eosin staining revealed that lipid deposition was focal in the TIS21-KO liver as opposed to the diffuse and homogeneous pattern in the WT liver after 24 h starvation. In addition, cathepsin E expression was over 10 times higher in the TIS21-KO liver than that in the WT, as opposed to the significant reduction of thioltransferase in both adult and fetal livers. At present, we cannot account for the role of cathepsin E. However, downregulation of glutaredoxin 2 thioltransferase expression might affect hypoxic damage in the TIS21-KO liver. We suggest that the $TIS21^{/BTG2}$ gene might be essential to maintain energy metabolism and reducing power in the liver under fasting condition.

• Molecules and Cells
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• v.41 no.7
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• pp.695-702
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• 2018

The inner ear is a complex sensory organ responsible for hearing and balance. Formation of the inner ear is dependent on tight regulation of spatial and temporal expression of genes that direct a series of developmental processes. Recently, epigenetic regulation has emerged as a crucial regulator of the development of various organs. However, what roles higher-order chromatin organization and its regulator molecules play in inner ear development are unclear. CCCTC-binding factor (CTCF) is a highly conserved 11-zinc finger protein that regulates the three-dimensional architecture of chromatin, and is involved in various gene regulation processes. To delineate the role of CTCF in inner ear development, the present study investigated inner ear-specific Ctcf knockout mouse embryos (Pax2-Cre; $Ctcf^{fl/fl}$). The loss of Ctcf resulted in multiple defects of inner ear development and severely compromised otic neurogenesis, which was partly due to a loss of Neurog1 expression. Furthermore, reduced Neurog1 gene expression by CTCF knockdown was found to be associated with changes in histone modification at the gene's promoter, as well as its upstream enhancer. The results of the present study demonstrate that CTCF plays an essential role in otic neurogenesis by modulating histone modification in the Neurog1 locus.

• Biotechnology and Bioprocess Engineering:BBE
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• v.11 no.6
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• pp.530-537
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• 2006

Recently isolated, Pseudomonas putida SN1 grows on styrene as its sole carbon and energy source through successive oxidation of styrene by styrene monooxygenase (SMO), styrene oxide isomerase (SOI), and phenylacetaldehyde dehydrogenase. For the production of (S)-styrene oxide, two knockout mutants of SN1 were constructed, one lacking SOI and another lacking both SMO and SOI. These mutants were developed into whole-cell biocatalysts by transformation with a multicopy plasmid vector containing SMO genes (styAB) of the SN1. Neither of these self-cloned recombinants could grow on styrene, but both converted styrene into an enantiopure (S)-styrene oxide (e.e. > 99%). Whole-cell SMO activity was higher in the recombinant constructed from the SOI-deleted mutant (130 U/g cdw) than in the other one (35 U/g cdw). However, the SMO activity of the former was about the same as that of the SOI-deleted SN1 possessing a single copy of the styAB gene that was used as host. This indicates that the copy number of styAB genes is not rate-limiting on SMO catalysis by whole-cell SN1.

• Journal of Microbiology and Biotechnology
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• v.16 no.10
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• pp.1622-1628
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• 2006

Bacteria from the Methylobacterium genus, called pink-pigmented facultative methylotrophic bacteria (PPFMs), are common inhabitants of plants, potentially dominating the phyllosphere population, and are also encountered in the rhizosphere, seeds, and other parts of plants, being versatile in nature. The consistent success of the Methylobacterium plant association relies on methylotrophy, the ability to utilize the one-carbon compound methanol emitted by plants. However, the efficiency of Methylobacterium in plant growth promotion could be better exploited and thus has attracted increasing interest in recent years. Accordingly, the present study investigated the inoculation effects of Methylobacterium sp. strains CBMB20 and CBMB 110 on seed imbibition to tomato and red pepper on the growth and accumulation of phytohormone levels under gnotobiotic conditions. Seeds treated with the Methylobacterium strains showed a significant increase in root length when compared with either the uninoculated control or Methylobacterium extorquens $miaA^-$ knockout mutanttreated seeds. Extracts of the plant samples were used for indole-3-acetic acid (IAA), trans-zeatin riboside (t-ZR), and dihydrozeatin riboside (DHZR) assays by immunoanalysis. The treatment with Methylobacterium sp. CBMB20 or CBMB 110 produced significant increases in the accumulation of IAA and the cytokinins t-ZR and DHZR in the red pepper extracts, whereas no IAA was detected in the tomato extracts, although the cytokinin concentrations were significantly increased. Therefore, this study proved that the versatility of Methylobacterium as a plant-growth promoting bacteria could be better exploited.