Faculty Profiles - NAKAMURA Yoshimasa

Papers - NAKAMURA Yoshimasa

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Mechanisms of the Selenium Tolerance of the Arabidopsis thaliana Knockout Mutant of Sulfate Transporter SULTR1;2 Reviewed

Misao Ohno, Misugi Uraji, Yasuaki Shimoishi, Izumi C. Mori, Yoshimasa Nakamura, Yoshiyuki Murata

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 76 ( 5 ) 993 - 998 2012.5

Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

We investigated the mechanism of selenium (Se) tolerance using an Arabidopsis thaliana knockout mutant of a sulfate transporter, sultr1;2. Se stress inhibited plant growth, decreased chlorophyll contents, and increased protein oxidation and lipid peroxidation in the wild type, whereas the sultr1;2 mutation mitigated damage of these forms, indicating that sultr1;2 is more tolerant of Se than the wild type is. The accumulation of symplastic Se was suppressed in sultr1;2 as compared to the wild type, and the chemical speciation of Se in the mutant was different from that in the wild type. Regardless of Se stress, the activities of ascorbate peroxidase, catalase, and peroxidase in the mutant were higher than in the wild type, while the activity of superoxide dismutase in the mutant was the same as in the wild type. These results suggest that the sultr1;2 mutation confers Se tolerance on Arabidopsis by decreasing symplastic Se and maintaining antioxidant enzyme activities.

DOI： 10.1271/bbb.111000

Effect of gamma Irradiation on the Fatty Acid Composition of Soybean and Soybean Oil Reviewed

Ikuko Minami, Yoshimasa Nakamura, Setsuko Todoriki, Yoshiyuki Murata

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 76 ( 5 ) 900 - 905 2012.5

Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

Food irradiation is a form of food processing to extend the shelf life and reduce spoilage of food. We examined the effects of gamma radiation on the fatty acid composition, lipid peroxidation level, and antioxidative activity of soybean and soybean oil which both contain a large amount of unsaturated fatty acids. Irradiation at 10 to 80 kGy under aerobic conditions did not markedly change the fatty acid composition of soybean. While 10-kGy irradiation did not markedly affect the fatty acid composition of soybean oil under either aerobic or anaerobic conditions, 40-kGy irradiation considerably altered the fatty acid composition of soybean oil under aerobic conditions, but not under anaerobic conditions. Moreover, 40-kGy irradiation produced a significant amount of trans fatty acids under aerobic conditions, but not under anaerobic conditions. Irradiating soybean oil induced lipid peroxidation and reduced the radical scavenging activity under aerobic conditions, but had no effect under anaerobic conditions. These results indicate that the fatty acid composition of soybean was not markedly affected by radiation at 10 kGy, and that anaerobic conditions reduced the degradation of soybean oil that occurred with high doses of gamma radiation.

DOI： 10.1271/bbb.110859

Cooperative Function of PLD delta and PLD alpha 1 in Abscisic Acid-Induced Stomatal Closure in Arabidopsis Reviewed

Misugi Uraji, Takeshi Katagiri, Eiji Okuma, Wenxiu Ye, Mohammad Anowar Hossain, Choji Masuda, Aya Miura, Yoshimasa Nakamura, Izumi C. Mori, Kazuo Shinozaki, Yoshiyuki Murata

PLANT PHYSIOLOGY 159 ( 1 ) 450 - 460 2012.5

Language：English Publishing type：Research paper (scientific journal) Publisher：AMER SOC PLANT BIOLOGISTS

Phospholipase D (PLD) is involved in responses to abiotic stress and abscisic acid (ABA) signaling. To investigate the roles of two Arabidopsis (Arabidopsis thaliana) PLDs, PLD alpha 1 and PLD delta, in ABA signaling in guard cells, we analyzed ABA responses in guard cells using Arabidopsis wild type, pld alpha 1 and pld delta single mutants, and a pld alpha 1 pld delta double mutant. ABA-induced stomatal closure was suppressed in the pld alpha 1 pld delta double mutant but not in the pld single mutants. The pld alpha 1 and pld delta mutations reduced ABA-induced phosphatidic acid production in epidermal tissues. Expression of either PLD alpha 1 or PLD delta complemented the double mutant stomatal phenotype. ABA-induced stomatal closure in both pld alpha 1 and pld delta single mutants was inhibited by a PLD inhibitor (1-butanol), suggesting that both PLD alpha 1 and PLD delta function in ABA-induced stomatal closure. During ABA-induced stomatal closure, wild-type guard cells accumulate reactive oxygen species and nitric oxide and undergo cytosolic alkalization, but these changes are reduced in guard cells of the pld alpha 1 pld delta double mutant. Inward-rectifying K+ channel currents of guard cells were inhibited by ABA in the wild type but not in the pld alpha 1 pld delta double mutant. ABA inhibited stomatal opening in the wild type and the pld delta mutant but not in the pld alpha 1 mutant. In wild-type rosette leaves, ABA significantly increased PLD delta transcript levels but did not change PLD alpha 1 transcript levels. Furthermore, the pld alpha 1 and pld delta mutations mitigated ABA inhibition of seed germination. These results suggest that PLD alpha 1 and PLD delta cooperate in ABA signaling in guard cells but that their functions do not completely overlap.

DOI： 10.1104/pp.112.195578

Luteolin Overcomes Resistance to Benzyl Isothiocyanate-Induced Apoptosis in Human Colorectal Cancer HCT-116 Cells Invited Reviewed

Rieko Sakai, Shintaro Yokobe, Naomi Abe, Noriyuki Miyoshi, Yoshiyuki Murata, Yoshimasa Nakamura

JOURNAL OF FOOD AND DRUG ANALYSIS 20 389 - 393 2012.4

Authorship：Last author,　Corresponding author Language：English Publishing type：Research paper (scientific journal) Publisher：FOOD & DRUG ADMINSTRATION

We have previously identified p53, a universal sensor of genotoxic stress, as a negative regulator of the apoptosis induction by benzyl isothiocyanate (BITC) in the normal colon fibroblastoid cells. In the present study, we further confirmed that BITC has a potential to induce cytotoxicity in the p53-mutated colon cancer HT-29 cells in preference to HCT-116 cells with wild-type p53. To obtain effective induction of BITC-stimulated apoptosis in p53-positive cells, we investigated the combination effect of BITC and food ingredient that may overcome resistance to BITC. Pretreatment with luteolin potentiated the cytotoxicity induction by BITC in HCT-116 cells but not in HT-29 cells. The biochemical events related to apoptosis such as DNA ladder formation and caspase-3 activation were also enhanced by luteolin. Luteolin attenuated the expression of p21(wafl/cipl) a key downstream target of p53. These results suggest the role of p21(wafl/cipl) pathway in the overcoming BITC resistance by luteolin.

Increased halogenated tyrosine levels are useful markers of human skin ageing, reflecting proteins denatured by past skin inflammation Reviewed

Y. Ishitsuka, F. Maniwa, C. Koide, Y. Kato, Y. Nakamura, T. Osawa, M. Tanioka, Y. Miyachi

CLINICAL AND EXPERIMENTAL DERMATOLOGY 37 ( 3 ) 252 - 258 2012.4

Language：English Publishing type：Research paper (scientific journal) Publisher：WILEY-BLACKWELL

Background. Photoageing of skin is thought to be caused by protein denaturation, which can be induced by ultraviolet radiation. Previous studies have also reported that inflammation is related to protein denaturation; however, the influence of inflammation on skin ageing has not been explored in detail.
Aim. To investigate the possible connection between inflammation and protein denaturation, which might lead to skin ageing, we focused on halogenated tyrosine as a denatured substance produced during the inflammation process.
Methods. We measured halogenated tyrosine in aged human skin. Inflammatory cells and halogenated tyrosine were detected by immunohistochemistry using antibodies to mast-cell tryptase, neutrophilic myeloperoxidase and halogenated tyrosine. Finally, using elastic van Gieson ( EVG) staining, we investigated whether the sites of halogenated tyrosine coincided with the sites at which proteins were denatured.
Results. Immunohistochemical analysis indicated that both inflammatory cells and halogenated tyrosines increased with ageing in both photoexposed and photoprotected skin. EVG staining confirmed that the localization of halogenated tyrosine was close to the sites at which protein was denatured.
Conclusions. Our investigations indicate a possible connection between skin ageing and inflammation, suggesting that halogenated tyrosine could be a useful marker of ageing skin.

DOI： 10.1111/j.1365-2230.2011.04215.x

Inhibitory Effects of Methylglyoxal on Light-Induced Stomatal Opening and Inward K+ Channel Activity in Arabidopsis Reviewed

Tahsina Sharmin Hoque, Eiji Okuma, Misugi Uraji, Takuya Furuichi, Takayuki Sasaki, Md Anamul Hoque, Yoshimasa Nakamura, Yoshiyuki Murata

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 76 ( 3 ) 617 - 619 2012.3

Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

Methylglyoxal (MG) is a reactive aldehyde derived by glycolysis. In Arabidopsis, MG inhibited light-induced stomatal opening in a dose-dependent manner. It significantly inhibited both inward-rectifying potassium (K-in) channels in guard-cell protoplasts and an Arabidopsis K-in channel, KAT1, heterologously expressed in Xenopus oocytes. Thus it appears that MG inhibition of stomatal opening involves MG inhibition of K+ influx into guard cells.

DOI： 10.1271/bbb.110885

FIA functions as an early signal component of abscisic acid signal cascade in Vicia faba guard cells Reviewed

Yusuke Sugiyama, Misugi Uraji, Megumi Watanabe-Sugimoto, Eiji Okuma, Shintaro Munemasa, Yasuaki Shimoishi, Yoshimasa Nakamura, Izumi C. Mori, Sumio Iwai, Yoshiyuki Murata

JOURNAL OF EXPERIMENTAL BOTANY 63 ( 3 ) 1357 - 1365 2012.2

Language：English Publishing type：Research paper (scientific journal) Publisher：OXFORD UNIV PRESS

An abscisic acid (ABA)-insensitive Vicia faba mutant, fia (fava bean impaired in ABA-induced stomatal closure) had previously been isolated. In this study, it was investigated how FIA functions in ABA signalling in guard cells of Vicia faba. Unlike ABA, methyl jasmonate (MeJA), H2O2, and nitric oxide (NO) induced stomatal closure in the fia mutant. ABA did not induce production of either reactive oxygen species or NO in the mutant. Moreover, ABA did not suppress inward-rectifying K+ (K-in) currents or activate ABA-activated protein kinase (AAPK) in mutant guard cells. These results suggest that FIA functions as an early signal component upstream of AAPK activation in ABA signalling but does not function in MeJA signalling in guard cells of Vicia faba.

DOI： 10.1093/jxb/err369

alpha-Tocopherol Sensitizes Human Leukemia HL-60 Cells to Apoptosis Induced by Benzyl Isothiocyanate Reviewed

Naomi Abe, Tomomi Shimizu, Noriyuki Miyoshi, Yoshiyuki Murata, Yoshimasa Nakamura

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 76 ( 2 ) 381 - 383 2012.2

Authorship：Last author,　Corresponding author Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

Pre-treatment with alpha-tocopherol (alpha-Toc) potentiated cytotoxicity induction by benzyl isothiocyanate (BITC). Biochemical events related to apoptosis, such as DNA ladder formation and caspase-3 activation, were also enhanced by alpha-Toc. These results suggest a significant role of the caspase-3 pathway in apoptosis induction regulated by alpha-Toc in combination.

DOI： 10.1271/bbb.110665

Cytotoxicity of Benzyl Isothiocyanate in Normal Renal Proximal Tubular Cells and Its Modulation by Glutathione Reviewed

Naomi Abe, Masashi Okuhira, Chiharu Tsutsui, Yoshiyuki Murata, Yoshimasa Nakamura

JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 60 ( 7 ) 1887 - 1892 2012.2

Authorship：Last author,　Corresponding author Language：English Publishing type：Research paper (scientific journal) Publisher：AMER CHEMICAL SOC

In the present study, we examined the toxicity of benzyl 1TC (BITC) and its urinary mercapturic acid metabolite (BITC-NAC), using a normal renal proximal tubular cell line, pig LLC-PK1. BITC increased cell death with an IC50 value of about 7 mu M, whereas the cytotoxic effect of BITC-NAC was five times weaker than that of BITC. We observed a significant necrosis of the compounds on LLC-PK1 cells with oxidative stress. In the presence of 5 mM glutathione (GSH), comparable to physiological levels, the cytotoxicity of BITC-NAC as well as BITC was significantly reduced. Furthermore, the increase in intracellular GSH levels by pretreatment with NAC before the BITC treatment resulted ill inhibition of the BITC-induced necrotic events as well as intracellular oxidative stress. These results suggest that GSH is a determinant of cellular resistance against the BITC-mediated and oxidative stress-dependent cytotoxicity in renal proximal tubular cells.

DOI： 10.1021/jf2052042

K252a-sensitive protein kinases but not okadaic acid-sensitive protein phosphatases regulate methyl jasmonate-induced cytosolic Ca2+ oscillation in guard cells of Arabidopsis thaliana Reviewed

Mohammad Anowar Hossain, Shintaro Munemasa, Yoshimasa Nakamura, Izumi C. Mori, Yoshiyuki Murata

JOURNAL OF PLANT PHYSIOLOGY 168 ( 16 ) 1901 - 1908 2011.11

Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER GMBH, URBAN & FISCHER VERLAG

Methyl jasmonate (MeJA) induces stomatal closure similar to abscisic acid (ABA), and MeJA signaling in guard cells shares some signal components with ABA signaling. As part of this process, MeJA as well as ABA induce the elevation and oscillation of cytosolic free-calcium concentrations ([Ca2+](cyt)) in guard cells. While abscisic acid-induced [Ca2+](cyt) oscillation has been extensively studied, MeJA-induced [Ca2+](cyt) oscillation is less well understood. In this study, we investigated the effects of K252a (a broad-range protein kinase inhibitor) and okadaic acid (OA, a protein phosphatase 1 and 2A inhibitor) on MeJA-induced [Ca2+](cyt) oscillation in guard cells of Arabidopsis thaliana ecotype Columbia expressing the Ca2+ reporter yellow cameleon 3.6. The protein kinase inhibitor K252a abolished MeJA-induced stomatal closure and reduced MeJA-elicited [Ca2+](cyt) oscillation. The protein phosphatase inhibitor OA, on the other hand, did not inhibit these processes. These results suggest that MeJA signaling involves activation of K252a-sensitive protein kinases upstream of [Ca2+](cyt) oscillation but not activation of an OA-sensitive protein phosphatase in guard cells of A. thaliana ecotype Columbia. (C) 2011 Elsevier GmbH. All rights reserved.

DOI： 10.1016/j.jplph.2011.05.004

Allyl isothiocyanate (AITC) induces stomatal closure in Arabidopsis Reviewed

Md. Atiqur Rahman Khokon, Md. Sarwar Jahan, Taniya Rahman, Mohammad Anowar Hossain, Daichi Muroyama, Ikuko Minami, Shintaro Munemasa, Izumi C. Mori, Yoshimasa Nakamura, Yoshiyuki Murata

PLANT CELL AND ENVIRONMENT 34 ( 11 ) 1900 - 1906 2011.11

Language：English Publishing type：Research paper (scientific journal) Publisher：WILEY-BLACKWELL

Isothiocyanates (ITCs) are degradation products of glucosinolates in crucifer plants and have repellent effect on insects, pathogens and herbivores. In this study, we report that exogenously applied allyl isothiocyanate (AITC) induced stomatal closure in Arabidopsis via production of reactive oxygen species (ROS) and nitric oxide (NO), and elevation of cytosolic Ca2+. AITC-induced stomatal closures were partially inhibited by an inhibitor of NADPH oxidase and completely inhibited by glutathione monoethyl ester (GSHmee). AITC-induced stomatal closure and ROS production were examined in abscisic acid (ABA) deficient mutant aba2-2 and methyl jasmonate (MeJA)-deficient mutant aos to elucidate involvement of endogenous ABA and MeJA. Genetic evidences have demonstrated that AITC-induced stomatal closure required MeJA priming but not ABA priming. These results raise the possibility that crucifer plants produce ITCs to induce stomatal closure, leading to suppression of water loss and invasion of fungi through stomata.

DOI： 10.1111/j.1365-3040.2011.02385.x

Roles of intracellular hydrogen peroxide accumulation in abscisic acid signaling in Arabidopsis guard cells Reviewed

Rayhanur Jannat, Misugi Uraji, Miho Morofuji, Mohammad Muzahidul Islam, Rachel E. Bloom, Yoshimasa Nakamura, C. Robertson McClung, Julian I. Schroeder, Izumi C. Mori, Yoshiyuki Murata

JOURNAL OF PLANT PHYSIOLOGY 168 ( 16 ) 1919 - 1926 2011.11

Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER GMBH, URBAN & FISCHER VERLAG

Reactive oxygen species (ROS), including hydrogen peroxide (H(2)O(2)), are among the important second messengers in abscisic acid (ABA) signaling in guard cells. In this study, to investigate specific roles of H(2)O(2) in ABA signaling in guard cells, we examined the effects of mutations in the guard cell-expressed catalase (CAT) genes, CAT1 and CAT3, and of the CAT inhibitor 3-aminotriazole (AT) on stomata! movement. The cat3 and cat1 cat3 mutations significantly reduced CAT activities, leading to higher basal level of H(2)O(2) in guard cells, when assessed by 2',7'-dichlorodihydrofluorescein, whereas they did not affect stomatal aperture size under non-stressed condition. In addition, AT-treatment at concentrations that abolish CAT activities, showed trivial affect on stomatal aperture size, while basal H(2)O(2) level increased extensively. In contrast, cat mutations and AT-treatment potentiated ABA-induced stomata! closure. Inducible ROS production triggered by ABA was observed in these mutants and wild type as well as in AT-treated guard cells. These results suggest that ABA-inducible cytosolic H(2)O(2) elevation functions in ABA-induced stomatal closure, while constitutive increase of H(2)O(2) do not cause stomatal closure. (C) 2011 Elsevier GmbH. All rights reserved.

DOI： 10.1016/j.jplph.2011.05.006

Negative regulation of abscisic acid-induced stomatal closure by glutathione in Arabidopsis Reviewed

Eiji Okuma, Md Sarwar Jahan, Shintaro Munemasa, Mohammad Anowar Hossain, Daichi Muroyama, Mohammad Mahbub Islam, Ken'ichi Ogawa, Megumi Watanabe-Sugimoto, Yoshimasa Nakamura, Yasuaki Shimoishi, Izumi C. Mori, Yoshiyuki Murata

JOURNAL OF PLANT PHYSIOLOGY 168 ( 17 ) 2048 - 2055 2011.11

Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER GMBH, URBAN & FISCHER VERLAG

We found that glutathione (GSH) is involved in abscisic acid (ABA)-induced stomatal closure. Regulation of ABA signaling by GSH in guard cells was investigated using an Arabidopsis mutant. cad2-1. that is deficient in the first GSH biosynthesis enzyme, gamma-glutamylcysteine synthetase, and a GSH-decreasing chemical, 1-chloro-2,4-dinitrobenzene (CDNB). Glutathione contents in guard cells decreased along with ABA-induced stomata! closure. Decreasing GSH by both the cad2-1 mutation and CDNB treatment enhanced ABA-induced stomata! closure. Glutathione monoethyl ester (GSHmee) restored the GSH level in cad2-1 guard cells and complemented the stomatal phenotype of the mutant. Depletion of GSH did not significantly increase ABA-induced production of reactive oxygen species in guard cells and GSH did not affect either activation of plasma membrane Ca2+-permeable channel currents by ABA or oscillation of the cytosolic free Ca2+ concentration induced by ABA. These results indicate that GSH negatively modulates a signal component other than ROS production and Ca2+ oscillation in ABA signal pathway of Arabidopsis guard cells. (C) 2011 Elsevier GmbH. All rights reserved.

DOI： 10.1016/j.jplph.2011.06.002

Localized cysteine sulfenic acid formation by vascular endothelial growth factor: role in endothelial cell migration and angiogenesis Reviewed

Nihal Kaplan, Norifumi Urao, Eiji Furuta, Seok-Jo Kim, Masooma Razvi, Yoshimasa Nakamura, Ronald D. McKinney, Leslie B. Poole, Tohru Fukai, Masuko Ushio-Fukai

FREE RADICAL RESEARCH 45 ( 10 ) 1124 - 1135 2011.10

Language：English Publishing type：Research paper (scientific journal) Publisher：INFORMA HEALTHCARE

Reactive oxygen species (ROS) are important mediators for VEGF receptor 2 (VEGFR2) signalling involved in angiogenesis. The initial product of Cys oxidation, cysteine sulfenic acid (Cys-OH), is a key intermediate in redox signal transduction; however, its role in VEGF signalling is unknown. We have previously demonstrated IQGAP1 as a VEGFR2 binding scaffold protein involved in ROS-dependent EC migration and post-ischemic angiogenesis. Using a biotin-labelled Cys-OH trapping reagent, we show that VEGF increases protein-Cys-OH formation at the lamellipodial leading edge where it co-localizes with NADPH oxidase and IQGAP1 in migrating ECs, which is prevented by IQGAP1 siRNA or trapping of Cys-OH with dimedone. VEGF increases IQGAP1-Cys-OH formation, which is prevented by N-acetyl cysteine or dimedone, which inhibits VEGF-induced EC migration and capillary network formation. In vivo, hindlimb ischemia in mice increases CysOH formation in small vessels and IQGAP1 in ischemic tissues. In summary, VEGF stimulates localized formation of Cys-OH-IQGAP1 at the leading edge, thereby promoting directional EC migration, which may contribute to post-natal angiogenesis in vivo. Thus, targeting Cys-oxidized proteins at specific compartments may be the potential therapeutic strategy for various angiogenesis-dependent diseases.

DOI： 10.3109/10715762.2011.602073

The Roles of CATALASE2 in Abscisic Acid Signaling in Arabidopsis Guard Cells Reviewed

Rayhanur Jannat, Misugi Uraji, Miho Morofuji, Mohammad Anowar Hossain, Mohammad Muzahidul Islam, Yoshimasa Nakamura, Izumi C. Mori, Yoshiyuki Murata

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 75 ( 10 ) 2034 - 2036 2011.10

Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

We investigated the roles of catalase (CAT) in abscisic acid (ABA)-induced stomatal closure using a cat2 mutant and an inhibitor of CAT, 3-aminotriazole (AT). Constitutive reactive oxygen species (ROS) accumulation due to the CAT2 mutation and AT treatment did not affect stomatal aperture in the absence of ABA, whereas ABA-induced stomatal closure, ROS production, and [Ca2+](cyt) oscillation were enhanced.

DOI： 10.1271/bbb.110344

Histochemical quantification of glutathione contents in guard cells of Arabidopsis thaliana Reviewed

Md. Sarwar Jahan, Yoshimasa Nakamura, Yoshiyuki Murata

SCIENCEASIA 37 ( 3 ) 281 - 284 2011.9

Language：English Publishing type：Research paper (scientific journal) Publisher：THAILANDS NATL SCIENCE & TECHNOLOGY DEVELOPMENT AGENCY

Glutathione (GSH), one of the most abundant low molecular-weight thiol compounds, maintains redox homoeostasis in plants. To measure GSH content in guard cells of Arabidopsis thaliana, we have developed a technique to quantify the GSH content in guard cells using the staining dye monochlorobimane (MCB). The fluorescent intensity of glutathione S-bimane (GSB) gradually increased in guard cells with increasing incubation time and reached steady state 2 h after epidermal tissues were treated with MCB. The guard cells contained larger amount of GSH content than other cells of the leaves except trichome cell. MCB staining showed that guard cells of a GSH deficient mutant, accumulated significantly lower GSH content than that of wild type guard cells. When the guard cells were treated with 1-chloro-2,4-dinitrobenzene to reduce GSH content in the guard cells, the GSB fluorescence in the guard cells was abolished. In addition, GSH monoethyl ester restored GSH level in the guard cells of ch1-1 mutant plants. These results suggest that GSH content in guard cells can be quantified using MCB dye.

DOI： 10.2306/scienceasia1513-1874.2011.37.281

Hydogen peroxide-dependent photocytotoxicity by phloxine B, a xanthene-type food colorant Reviewed

Hang Qi, Hiroshi Takano, Yoji Kato, Qian Wu, Chiharu Ogata, Beiwei Zhu, Yoshiyuki Murata, Yoshimasa Nakamura

BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS 1810 ( 7 ) 704 - 712 2011.7

Authorship：Last author,　Corresponding author Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER SCIENCE BV

Background: Phloxine B (PhB; 2',4',5',7'-tetrabromo-4,5,6,7-tetrachloro-fluorescein), an artificial xanthene colorant, has been used as a red coloring agent in drugs and cosmetics as well as foods in some countries. However, little effort has been devoted to the study of this colorant as a potentially useful medicinal agent.
Methods: We investigated the daily light-induced photocytotoxicity of PhB in two human leukemia cells, HL-60 and Jurkat, and its underlying mechanisms by in vitro experiments using antioxidants.
Reuslts and conclusions: PhB inhibited cell proliferation more preferentially to HL-60 cells than to Jurkat cells. Co-treatment of catalase completely blocked the photocytotoxicity by PhB in HL-60 cells, whereas the effect of histidine was only partial, suggesting that hydrogen peroxide (H(2)O(2)), rather than singlet oxygen, might be a prerequisite for the PhB-induced HL-60 cell death. Actually, PhB produced a significant amount of H(2)O(2) in the media as well as in the cells in concentration- and light-dependent manners. Furthermore, methionine, a hypochlorous acid (HOCl) scavenger, also significantly attenuated the cytotoxicity in HL-60 cells, but not in Jurkat cells, indicating the involvement of myeloperoxidase (MPO)-dependent hypohalous acid formation during the photocytotoxicity. In vitro experiments revealed that halogenated tyrosine was generated from the reaction of bovine serum albumin with PhB and HL-60 cell lysate. The present findings suggested that PhB induced a differential photodynamic action in the MPO-containing leukemia cells through an H(2)O(2)-dependent mechanism.
General significance: Our findings provide new insights into the molecular mechanisms underlying the PhB-induced apoptosis and also evaluated PhB as a promising PDT agent. (C) 2011 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.bbagen.2011.04.010

Photostability of Lycopene Dispersed in an Aqueous Solution Reviewed

Masayuki Nishino, Takeshi Miuchi, Makoto Sakata, Atsushi Nishida, Yoshiyuki Murata, Yoshimasa Nakamura

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 75 ( 7 ) 1389 - 1391 2011.7

Authorship：Last author Language：English Publishing type：Research paper (scientific journal) Publisher：TAYLOR & FRANCIS LTD

Lycopene dispersed in aqueous solutions with different dissolved oxygen contents was photo-irradiated by using a xenon weather meter, and the contents of lycopene and dissolved oxygen were measured. Both the degradation of lycopene and the consumption of dissolved oxygen followed a first-order kinetics model. There was a proportional relationship between the degradation content of lycopene and the consumption of dissolved oxygen. These results indicate that dissolved oxygen would also be involved in the photolysis of lycopene.

DOI： 10.1271/bbb.110154

Involvement of Endogenous Abscisic Acid in Methyl Jasmonate-Induced Stomatal Closure in Arabidopsis Reviewed

Mohammad Anowar Hossain, Shintaro Munemasa, Misugi Uraji, Yoshimasa Nakamura, Izumi C. Mori, Yoshiyuki Murata

PLANT PHYSIOLOGY 156 ( 1 ) 430 - 438 2011.5

Language：English Publishing type：Research paper (scientific journal) Publisher：AMER SOC PLANT BIOLOGISTS

In this study, we examined the involvement of endogenous abscisic acid (ABA) in methyl jasmonate (MeJA)-induced stomatal closure using an inhibitor of ABA biosynthesis, fluridon (FLU), and an ABA-deficient Arabidopsis (Arabidopsis thaliana) mutant, aba2-2. We found that pretreatment with FLU inhibited MeJA-induced stomatal closure but not ABA-induced stomatal closure in wild-type plants. The aba2-2 mutation impaired MeJA-induced stomatal closure but not ABA-induced stomatal closure. We also investigated the effects of FLU and the aba2-2 mutation on cytosolic free calcium concentration ([Ca2+](cyt)) in guard cells using a Ca2+-reporter fluorescent protein, Yellow Cameleon 3.6. In wild-type guard cells, FLU inhibited MeJA-induced [Ca2+](cyt) elevation but not ABA-induced [Ca2+](cyt) elevation. The aba2-2 mutation did not affect ABA-elicited [Ca2+](cyt) elevation but suppressed MeJA-induced [Ca2+](cyt) elevation. We also tested the effects of the aba2-2 mutation and FLU on the expression of MeJA-inducible VEGETATIVE STORAGE PROTEIN1 (VSP1). In the aba2-2 mutant, MeJA did not induce VSP1 expression. In wild-type leaves, FLU inhibited MeJA-induced VSP1 expression. Pretreatment with ABA at 0.1 mu M, which is not enough concentration to evoke ABA responses in the wild type, rescued the observed phenotypes of the aba2-2 mutant. Finally, we found that in wild-type leaves, MeJA stimulates the expression of 9-CIS-EPOXYCAROTENOID DIOXYGENASE3, which encodes a crucial enzyme in ABA biosynthesis. These results suggest that endogenous ABA could be involved in MeJA signal transduction and lead to stomatal closure in Arabidopsis guard cells.

DOI： 10.1104/pp.111.172254

(-)-Epigallocatechin-3-gallate suppresses growth of AZ521 human gastric cancer cells by targeting the DEAD-box RNA helicase p68 Reviewed

Tomoko Tanaka, Takeshi Ishii, Daisuke Mizuno, Taiki Mori, Ryoichi Yamaji, Yoshimasa Nakamura, Shigenori Kumazawa, Tsutomu Nakayama, Mitsugu Akagawa

FREE RADICAL BIOLOGY AND MEDICINE 50 ( 10 ) 1324 - 1335 2011.5

Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER SCIENCE INC

(-)-Epigallocatechin-3-gallate (EGCG), the most abundant and biologically active polyphenol in green tea, induces apoptosis and suppresses proliferation of cancer cells by modulating multiple signal transduction pathways. However, the fundamental mechanisms responsible for these cancer-preventive effects have not been clearly elucidated. Recently, we found that EGCG can covalently bind to cysteine residues in proteins through autoxidation and subsequently modulate protein function. In this study, we demonstrate the direct binding of EGCG to cellular proteins in AZ521 human gastric cancer cells by redox-cycle staining. We comprehensively explored the binding targets of EGCG from EGCG-treated A2521 cells by proteomics techniques combined with the boronate-affinity pull-down method. The DEAD-box RNA helicase p68, which is overexpressed in a variety of tumor cells and plays an important role in cancer development and progression, was identified as a novel EGCG-binding target. Exposure of AZ521 cells to EGCG lowered the p68 level dose dependently. The present findings show that EGCG inhibits AZ521 cell proliferation by preventing beta-catenin oncogenic signaling through proteasomal degradation of p68 and provide a new perspective on the molecular mechanism of EGCG action. (C) 2011 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.freeradbiomed.2011.01.024

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