Faculty Profiles - MUNEMASA Shintaro

Papers - MUNEMASA Shintaro

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Involvement of reactive carbonyl species in inhibition of germination and seedling growth by salt stress in rice Invited

Faruk Hossain Khan, Yoshitaka Nakashima, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshihiko Hirai, Md Anamul Hoque, Mohammad Saidur Rhaman, Shintaro Munemasa, Jun'ichi Mano, Yoshiyuki Murata

Bioscience, Biotechnology, and Biochemistry 2025.5

Dual role of cytosolic GSH in the ABA signaling pathway and plasma membrane ion channel regulation in guard cells of Vicia faba. International journal

Huifei Yin, Toshiyuki Nakamura, Yoshimasa Nakamura, Shintaro Munemasa, Yoshiyuki Murata

Journal of plant physiology 306 154447 - 154447 2025.2

Language：English Publishing type：Research paper (scientific journal)

Abscisic acid (ABA) induces stomatal closure in higher plants under drought stress. Glutathione (GSH) negatively regulates ABA-induced stomatal closure and reactive carbonyl species (RCS) play a role as signal mediators downstream of reactive oxygen species production in ABA signaling pathway in Arabidopsis thaliana. Activation of slow (S-type) anion channels and inhibition of inward-rectifying potassium ion (Kin+) channels in the plasma membrane are essential for ABA-induced stomatal closure. However, there is limited evidence regarding role of GSH in the activation of S-type anion channels and the inhibition of Kin+ channels. We used Vicia faba to clarify the regulation of these ion channels by GSH and RCS. Pretreatment of guard-cell protoplasts with the GSH-supplementing agent, glutathione monoethyl ester (GSHmee), suppressed the activation of S-type anion channels and the inactivation of Kin+ channels induced by ABA. The pretreatment with the RCS scavenger carnosine suppressed the activation of S-type anion channels and the inactivation of Kin+ channels by ABA. On patch clamping guard-cell protoplasts, the addition of GSH to the pipette (cytosolic) buffer decreased the S-type anion currents and increased the Kin+ currents. These results suggest that cytosolic GSH is involved in ABA-induced stomatal closure via negative regulation of ABA signaling and via direct regulation of ion channel activities in V. faba.

DOI： 10.1016/j.jplph.2025.154447

TGG1 and TGG2 mutations impair allyl isothiocyanate-mediated stomatal closure in Arabidopsis thaliana. International journal

Kadri Oumaima, Mohammad Shakhawat Hossain, Wenxiu Ye, Eiji Okuma, Mohammad Issak, Mohammad Mahbub Islam, Misugi Uraji, Yoshimasa Nakamura, Izumi C Mori, Shintaro Munemasa, Yoshiyuki Murata

Protoplasma 2025.2

Language：English Publishing type：Research paper (scientific journal)

Myrosinase, referred to as thioglucoside glucohydrolase (TGG), plays a crucial role in plant physiology through catalyzing the hydrolysis of glucosinolates into bioactive isothiocyanates. In Arabidopsis thaliana, the myrosinases TGG1 and TGG2 are essential for abscisic acid- and methyl jasmonate-induced stomata closure. Allyl isothiocyanate (AITC), one of myrosinase products, triggers stomatal closure in A. thaliana. We investigated stomatal responses to AITC to clarify the role of TGG1 and TGG2 in Arabidopsis guard-cell signaling. Allyl isothiocyanate at 50 μM and 100 μM induced stomatal closure in the tgg1 and tgg2 single mutants but not in the tgg1 tgg2 double mutant. Furthermore, AITC at 50 μM induced the production of reactive oxygen species and nitric oxide, cytosolic alkalization, and oscillations in cytosolic free calcium concentration in guard cells of both wild-type and mutant plants. These findings suggest that TGG1 and TGG2 are involved in AITC signaling pathway through interaction with signal component(s) downstream of these signaling events, which is not accompanied by hydrolysis of glucosinolates because of the difference in subcellular localization between enzymes (myrosinases) and substrates (glucosinolates).

DOI： 10.1007/s00709-025-02039-z

Reactive carbonyl species function downstream of reactive oxygen species in chitosan-induced stomatal closure. International journal

Israt Jahan, Md Moshiul Islam, Toshiyuki Nakamura, Yoshimasa Nakamura, Shintaro Munemasa, Jun'ichi Mano, Yoshiyuki Murata

Physiologia plantarum 177 ( 1 ) e70094 2025

Language：English Publishing type：Research paper (scientific journal)

An elicitor, chitosan (CHT), induces stomatal closure in plants, which is accompanied by salicylhydroxamic acid (SHAM)-sensitive peroxidases-mediated reactive oxygen species (ROS) production in guard cells. Reactive carbonyl species (RCS) function downstream of ROS in abscisic acid (ABA) and methyl jasmonate (MeJA) signalling in guard cells. However, the involvement of RCS in CHT-induced stomatal closure is still unknown. In this study, we used transgenic tobacco (Nicotiana tabacum) plants overexpressing Arabidopsis thaliana 2-alkenal reductase (AER-OE tobacco) and Arabidopsis wild-type (WT) plants to investigate whether RCS is involved in CHT-induced stomatal closure. Chitosan-induced stomatal closure was inhibited in the tobacco AER-OE plants. In the WT tobacco and Arabidopsis plants, CHT-induced stomatal closure was inhibited by RCS scavengers, carnosine and pyridoxamine. Chitosan significantly increased RCS production in the WT tobacco and Arabidopsis, but in the tobacco AER-OE plants, chitosan did not increase significantly RCS accumulation. Moreover, neither the application of RCS scavengers to both WT plants nor scavenging RCS by AER-OE affected the CHT-induced ROS accumulation. However, treatment with a peroxidase inhibitor, SHAM, significantly inhibited CHT-induced RCS accumulation in WT tobacco and Arabidopsis plants. Taken together, these results suggest that RCS acts downstream of ROS production in CHT signalling in guard cells of A. thaliana and N. tabacum.

DOI： 10.1111/ppl.70094

GUARD CELL HYDROGEN PEROXIDE-RESISTANT1 functions upstream of reactive carbonyl species production in Arabidopsis guard-cell abscisic acid signaling Reviewed

Oumayma Shaiek, Huifei Yin, Nodoka Uesako, Md Moshiul Islam, Mohammad Saidur Rhaman, Toshiyuki Nakamura, Yoshimasa Nakamura, Shintaro Munemasa, Jun'ichi Mano, Yoshiyuki Murata

Bioscience, Biotechnology, and Biochemistry 88 ( 12 ) 1403 - 1410 2024.9

Publishing type：Research paper (scientific journal) Publisher：Oxford University Press (OUP)

Abstract

GUARD CELL HYDROGEN PEROXIDE-RESISTANT1 (GHR1), a leucine-rich repeat receptor-like kinase, is involved in abscisic acid (ABA)-induced stomatal closure. We investigated the role of GHR1 in reactive oxygen species (ROS) signaling for ABA-induced stomatal closure. ABA induced ROS production in wild type (WT) and the ghr1 of Arabidopsis thaliana. Hydrogen peroxide induced stomatal closure, accompanying the generation of acrolein in guard cells. The reactive carbonyl species (RCS) scavengers inhibited the ABA- and H2O2-induced stomatal closure in WT. In the ghr1, H2O2 failed to induce acrolein production and stomatal closure while RCS induced stomatal closure. Thus, GHR1 functions downstream of ROS and is required for the generation of RCS in guard-cell ABA signaling. In the ghr1, Ca2+ induced stomatal closure but RCS did not activate ICa channels. The GHR1 may be also involved in a Ca2+-independent pathway for ABA-induced stomatal closure.

DOI： 10.1093/bbb/zbae135

Other Link： https://academic.oup.com/bbb/article-pdf/88/12/1403/60785758/zbae135.pdf

Quercetin Attenuates Acetaldehyde-Induced Cytotoxicity via the Heme Oxygenase-1-Dependent Antioxidant Mechanism in Hepatocytes Reviewed

Kexin Li, Minori Kidawara, Qiguang Chen, Shintaro Munemasa, Yoshiyuki Murata, Toshiyuki Nakamura, Yoshimasa Nakamura

International Journal of Molecular Sciences 25 ( 16 ) 9038 - 9038 2024.8

Publishing type：Research paper (scientific journal) Publisher：MDPI AG

It is still unclear whether or how quercetin influences the toxic events induced by acetaldehyde in hepatocytes, though quercetin has been reported to mitigate alcohol-induced mouse liver injury. In this study, we evaluated the modulating effect of quercetin on the cytotoxicity induced by acetaldehyde in mouse hepatoma Hepa1c1c7 cells, the frequently used cellular hepatocyte model. The pretreatment with quercetin significantly inhibited the cytotoxicity induced by acetaldehyde. The treatment with quercetin itself had an ability to enhance the total ALDH activity, as well as the ALDH1A1 and ALDH3A1 gene expressions. The acetaldehyde treatment significantly enhanced the intracellular reactive oxygen species (ROS) level, whereas the quercetin pretreatment dose-dependently inhibited it. Accordingly, the treatment with quercetin itself significantly up-regulated the representative intracellular antioxidant-related gene expressions, including heme oxygenase-1 (HO-1), glutamate-cysteine ligase, catalytic subunit (GCLC), and cystine/glutamate exchanger (xCT), that coincided with the enhancement of the total intracellular glutathione (GSH) level. Tin protoporphyrin IX (SNPP), a typical HO-1 inhibitor, restored the quercetin-induced reduction in the intracellular ROS level, whereas buthionine sulphoximine, a representative GSH biosynthesis inhibitor, did not. SNPP also cancelled the quercetin-induced cytoprotection against acetaldehyde. These results suggest that the low-molecular-weight antioxidants produced by the HO-1 enzymatic reaction are mainly attributable to quercetin-induced cytoprotection.

DOI： 10.3390/ijms25169038

The Metabolite of γ-Tocopherol, 2,7,8-Trimethyl-2-(2′-Carboxyethyl)-6-Hydroxychroman, Exerts Intracellular Antioxidant Activity via Up-Regulation of Heme Oxygenase-1 in Hepatocytes Invited Reviewed

Shosuke Aoyama, Tomoka Nishio, Daiki Moriya, Shintaro Munemasa, Yoshiyuki Murata, Yoshimasa Nakamura, Toshiyuki Nakamura

Nutraceuticals 4 ( 3 ) 409 - 416 2024.8

Publishing type：Research paper (scientific journal) Publisher：MDPI AG

γ-Tocopherol (γT) is the major form of vitamin E contained in plants and seed oils. Although it is readily metabolized in the liver, the function of the metabolites is not fully understood. This study investigated the antioxidant activities of the γT metabolite 2,7,8-trimethyl-2-(2′-carboxyethyl)-6-hydroxychroman (γCEHC) in comparison to its parent compound. The pretreatment of mouse hepatoma Hepa1c1c7 cells with γCEHC showed a cytoprotective effect on the hydrogen peroxide-induced cytotoxicity to a lesser extent than that of γT. A mechanistic investigation revealed that both γ-CEHC and γT significantly up-regulated the gene and protein expressions of heme oxygenase-1 (HO-1) via the promotion of the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2). Furthermore, the combination of γCEHC and γT significantly increased the gene and protein levels of HO-1 and the nuclear translocation of Nrf2, suggesting that it was an additive effect. Tin protoporphyrin IX (SnPP), a representative HO-1 inhibitor, significantly impaired the cytoprotection of γCEHC and γT against the hydrogen peroxide-induced cytotoxicity. These results suggested that not only γT but also its metabolite, γCEHC, are a promising cytoprotective factor against oxidative stress-induced cytotoxicity and that the cytoprotective effect is attributable to the cooperation of both compounds.

DOI： 10.3390/nutraceuticals4030024

Outward-rectifying potassium channels GORK and SKOR function in regulation of root growth under salt stress in Arabidopsis thaliana. Reviewed International journal

Hafsa Jahan Hiya, Yoshitaka Nakashima, Airi Takeuchi, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshiyuki Murata, Shintaro Munemasa

Journal of plant physiology 302 154322 - 154322 2024.8

Authorship：Corresponding author Language：English Publishing type：Research paper (scientific journal)

Plants often face high salinity as a significant environmental challenge with roots being the first responders to this stress. Maintaining K+/Na+ ratio within plant cells is crucial for survival, as the intracellular K+ level decreases and the intracellular Na+ level increases under saline conditions. However, knowledge about the molecular regulatory mechanisms of K+ loss in response to salt stress through outward-rectifying K+ channels in plants is largely unknown. In this study, we found that the Arabidopsis double mutant gorkskor, in which the GORK and SKOR genes are disrupted, showed an improved primary root growth under salt stress compared to wild-type (WT) and the gork and skor single-mutant plants. No significant differences in the sensitivity to mannitol stress between the WT and gorkskor mutant were observed. Accumulation of ROS induced by salt stress was reduced in the gorkskor roots. The gorkskor mutant seedlings had significantly higher K+ content, lower Na+ content, and a greater resultant K+/Na+ ratio than the WT under salt stress. Moreover, salt-stress-induced elevation of cytosolic free Ca2+ concentration was reduced in the gorkskor roots. Taken together, these results suggest that Arabidopsis Shaker-type outward-rectifying K+ channels GORK and SKOR may redundantly function in regulation of primary root growth under salt stress and are involved in not only the late-stage response (e.g. K+ leakage) but also the early response including ROS production and [Ca2+]cyt elevation.

DOI： 10.1016/j.jplph.2024.154322

Evaluation of quercetin as a potential cytoprotector against acetaldehyde using the cultured hepatocyte model with aldehyde dehydrogenase isozyme deficiency. International journal

Yuhang Xu, Takeshi Sawamoto, Ruitong Sun, Aki Ishikura, Shintaro Munemasa, Yoshiyuki Murata, Ayano Satoh, Akiko Matsumoto, Toshiyuki Nakamura, Yoshimasa Nakamura

Bioscience, biotechnology, and biochemistry 2024.7

Language：English Publishing type：Research paper (scientific journal)

Protective effect of quercetin against acetaldehyde was evaluated using the cultured hepatocyte models with aldehyde dehydrogenase (ALDH) isozyme deficiency (aldh2-kd and aldh1a1-kd). The quercetin-induced cytoprotection against acetaldehyde in the ALDH1A1-deficient mutant (aldh1a1-kd) was weaker than that in wild type. Furthermore, quercetin did not enhance the ALDH activity in aldh1a1-kd cells, suggesting that ALDH1A1 is involved in the quercetin-induced cytoprotection.

DOI： 10.1093/bbb/zbae100

Negative regulation of chitosan-induced stomatal closure by glutathione in Arabidopsis thaliana. Reviewed International journal

Israt Jahan, Shintaro Munemasa, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshiyuki Murata

Bioscience, biotechnology, and biochemistry 2024.5

Language：English Publishing type：Research paper (scientific journal)

Chitosan (CHT) is a deacylated derivative of chitin and improves growth and yield performance, activates defensive genes, and also induces stomatal closure in plants. Glutathione (GSH) has significant functions in the growth, development, defense systems, signaling, and gene expression. Glutathione negatively regulates abscisic acid (ABA)-, methyl jasmonate (MeJA)-, and salicylic acid (SA)-induced stomatal closure. However, the negative regulation by GSH of CHT-induced stomatal closure is still unknown. Regulation of CHT-induced stomatal closure by GSH in guard cells was investigated using two GSH-deficient mutants, cad2-1 and ch1-1, and a GSH-decreasing chemical, 1-chloro-2,4-dinitrobenzene (CDNB). The cad2-1 and ch1-1 mutations and CDNB treatment enhanced CHT-induced stomatal closure. Treatment with glutathione monoethyl ester (GSHmee) restored the GSH level in the guard cells of cad2-1 and ch1-1 and complemented the stomatal phenotype of the mutants. These results indicate that GSH negatively regulates CHT-induced stomatal closure in A. thaliana.

DOI： 10.1093/bbb/zbae065

Enhancing effect of the coexisting alpha-tocopherol on quercetin absorption and metabolism. International journal

Rikito Mitsuzane, Reiko Okubo, Miyu Nishikawa, Shinichi Ikushiro, Shintaro Munemasa, Yoshiyuki Murata, Yoshimasa Nakamura, Toshiyuki Nakamura

Free radical research 1 - 16 2024.2

Language：English Publishing type：Research paper (scientific journal)

The aim of this study is to investigate the modulating effect of coexisting food components on the absorption and metabolism of quercetin and blood plasma antioxidant potentials. The combination of quercetin with α-tocopherol (αT), cellulose, or a commercially-available vegetable beverage containing αT and dietary fiber was orally administered to mice. Compared to the single administration of quercetin aglycone, the co-administration of αT with quercetin significantly increased the plasma quercetin concentration at 0.5 h, whereas the combination of quercetin and cellulose decreased it. Interestingly, the administration of quercetin mixed with the vegetable beverage showed no significant change of the quercetin concentration in the mice plasma. The treatment of the cells with the blood plasma after the co-administration of αT with quercetin significantly upregulated the gene expression of the antioxidant enzyme (heme oxygenase-1), whereas the quercetin and cellulose combination did not. In the plasma of the quercetin-administered mice, eight types of quercetin metabolites were detected and their quantities were affected by the combination with αT. The potentials of the heme oxygenase-1 gene expression by these metabolites were very limited, although several metabolites showed radical scavenging activities comparable to aglycone in the in vitro assays. These results suggested that the combination of αT potentiates the quercetin absorption and metabolism and thus the plasma antioxidant potentials, at least in part, by the quantitative changes in the quercetin metabolites.

DOI： 10.1080/10715762.2024.2317206

An outward-rectifying plant K+ channel SPORK2 exhibits temperature-sensitive ion-transport activity. Reviewed International journal

Yuki Muraoka, Gangqiang Yang, Shintaro Munemasa, Yusuke Takeuchi, Yasuhiro Ishimaru, Yoshiyuki Murata, Nobuyuki Uozumi, Minoru Ueda

Current biology 33 5488 - 5494 2023.11

Language：English Publishing type：Research paper (scientific journal)

Temperature sensing is critical for the survival of living organisms.1,2 Thermosensitive transient receptor-potential (TRP) cation channels function as thermosensors in mammals.2,3,4,5,6 In contrast to animals, land plants lack TRP genes.7,8,9 Previous patch-clamp studies in plant cells suggested the presence of ion channels whose activities are related to temperature, implying the presence of TRP-like channels.10,11,12,13,14 However, the molecular entities of such temperature-sensitive ion channels were still unknown in land plants. In this study, we observed that the unique rainfall-induced leaf-folding movement of the legume tree Samanea saman15 was temperature-sensitive by using a rainfall-mimicking assay. Chilling-induced leaf folding in S. saman was shown to be related to the swelling of the motor cells16,17 at the base of the leaflet. This swelling suggested involvement of temperature-sensitive inactivation of K+ currents, independent of fluctuations in ion channel gene expression in motor cells. These findings led us to examine the temperature sensitivity of an outward-rectifying K+ channel, SPORK2, which was reported as an ion channel responsible for the nyctinastic (circadian-rhythmic) leaf movement of S. saman.18 We also discovered that SPORK2 exhibits temperature-sensitive K+ transport activity in the Xenopus oocyte expression system. Using chimeric channels, we showed that two domains of SPORK2 regulated the temperature sensitivity. Furthermore, heterologously expressed SPORK2 in Arabidopsis guard cells induced temperature-dependent stomatal closure. Therefore, SPORK2 is an ion channel in land plants with temperature-sensitive ion-transport activity that functions similarly to mammalian TRP channels. Our current findings advance the molecular understanding of temperature-sensing mechanisms in plants.

DOI： 10.1016/j.cub.2023.10.057

The effect of exogenous dihydroxyacetone and methylglyoxal on growth, anthocyanin accumulation, and the glyoxalase system in Arabidopsis. Reviewed International coauthorship International journal

Maoxiang Zhao, Toshiyuki Nakamura, Yoshimasa Nakamura, Shintaro Munemasa, Izumi C Mori, Yoshiyuki Murata

Bioscience, biotechnology, and biochemistry 87 1323 - 1331 2023.8

Language：English Publishing type：Research paper (scientific journal)

Dihydroxyacetone (DHA) occurs in wide-ranging organisms including plants and can undergo spontaneous conversion to methylglyoxal (MG). While the toxicity of MG to plants is well-known, the toxicity of DHA to plants remains to be elucidated. We investigated the effects of DHA and MG on Arabidopsis. Exogenous DHA at up to 10 m m did not affect the radicle emergence, the expansion of green cotyledons, the seedling growth, or the activity of glyoxalase II while DHA at 10 m m inhibited the root elongation and increased the activity of glyoxalase I. Exogenous MG at 1.0 m m inhibited these physiological responses and increased both activities. DHA at 10 m m increased the MG content in the roots. These results indicate that DHA is not so toxic as MG in Arabidopsis seeds and seedlings and suggest that the toxic effect of DHA at high concentrations is attributed to MG accumulation by the conversion to MG.

DOI： 10.1093/bbb/zbad109

Cycloartenyl Ferulate Is the Predominant Compound in Brown Rice Conferring Cytoprotective Potential against Oxidative Stress-Induced Cytotoxicity Reviewed International coauthorship International journal

Hongyan Wu, Toshiyuki Nakamura, Yingnan Guo, Riho Matsumoto, Shintaro Munemasa, Yoshiyuki Murata, Yoshimasa Nakamura

International Journal of Molecular Sciences 24 ( 1 ) 822 - 822 2023.1

Publishing type：Research paper (scientific journal) Publisher：MDPI AG

Since brown rice extract is a rich source of biologically active compounds, the present study is aimed to quantify the major compounds in brown rice and to compare their cytoprotective potential against oxidative stress. The content of the main hydrophobic compounds in brown rice followed the order of cycloartenyl ferulate (CAF) (89.00 ± 8.07 nmol/g) >> α-tocopherol (αT) (19.73 ± 2.28 nmol/g) > γ-tocotrienol (γT3) (18.24 ± 1.41 nmol/g) > α-tocotrienol (αT3) (16.02 ± 1.29 nmol/g) > γ-tocopherol (γT) (3.81 ± 0.40 nmol/g). However, the percent contribution of CAF to the radical scavenging activity of one gram of whole brown rice was similar to those of αT, αT3, and γT3 because of its weaker antioxidant activity. The CAF pretreatment displayed a significant cytoprotective effect on the hydrogen peroxide-induced cytotoxicity from 10 µM, which is lower than the minimal concentrations of αT and γT required for a significant protection. CAF also enhanced the nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation coincided with the enhancement of the heme oxygenase-1 (HO-1) mRNA level. An HO-1 inhibitor, tin protoporphyrin IX (SnPP), significantly impaired the cytoprotection of CAF. The cytoprotective potential of CAF is attributable to its cycloartenyl moiety besides the ferulyl moiety. These results suggested that CAF is the predominant cytoprotector in brown rice against hydrogen peroxide-induced cytotoxicity.

DOI： 10.3390/ijms24010822

The microbiota catabolites of quercetin glycosides concertedly enhance the resistance against acetaldehyde-induced oxidative stress. Reviewed International coauthorship International journal

Kexin Li, Hongyan Wu, Minori Kidawara, Yun Lin, Ayano Satoh, Gongliang Zhang, Shintaro Munemasa, Yoshiyuki Murata, Toshiyuki Nakamura, Yoshimasa Nakamura

Free radical research 56 1 - 14 2022.12

Language：English Publishing type：Research paper (scientific journal)

3,4-Dihydroxyphenylacetic acid (DOPAC) and 3-hydroxyphenylacetic acid (OPAC) are the predominant catabolites of quercetin glycosides, such as quercetin 4'-O-β-glucoside from the onion, produced by intestinal microbiota. Although each catabolite has been reported to protect the cells from acetaldehyde-induced cytotoxicity, the effect of their combination remains to be clarified. The purpose of this study was to determine whether the combination of DOPAC and OPAC enhances the resistance against the acetaldehyde-induced oxidative stress in the cultured hepatocytes. The pretreatment of the combination of DOPAC (5 μM) and OPAC (5 μM) showed the significant protection against the acetaldehyde- and hydrogen peroxide-induced cytotoxicity, even though each compound at the same concentration did not. This combination also significantly inhibited the intracellular dichlorofluorescin diacetate-detectable reactive oxygen species (ROS) level, whereas the solo treatment did slightly, suggesting that reducing mechanisms of ROS or compounds that enhance ROS production are involved in the cytoprotective effect. The combinatory treatment significantly enhanced the gene expression of not only the aldehyde dehydrogenases (ALDHs), but also glutamate-cysteine ligase, catalytic subunit, the first rate-limiting enzyme of glutathione (GSH) synthesis. Accordingly, both the intracellular GSH level and the total ALDH activity were enhanced by DOPAC plus OPAC. Involvement of GSH in the cytoprotection as well as ALDH up-regulation by the combination were confirmed by the experiments using a GSH biosynthesis inhibitor, buthionine sulfoximine. Taken together, the present results suggested that the quercetin microbiota catabolites concertedly protect the cells from acetaldehyde through a pre-enhanced resistance against oxidative stress by the GSH-dependent up-regulation of ALDHs.

DOI： 10.1080/10715762.2022.2159820

Extracellular malate induces stomatal closure via direct activation of guard‐cell anion channel SLAC1 and stimulation of Ca²⁺ signalling Reviewed International journal

Yoshiharu Mimata, Shintaro Munemasa, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshiyuki Murata

New Phytologist 236 ( 3 ) 852 - 863 2022.11

Malate induces stomatal closure via a receptor-like kinase GHR1- and reactive oxygen species-dependent pathway in Arabidopsis thaliana Reviewed International coauthorship International journal

Yoshiharu Mimata, Shintaro Munemasa, Fahmida Akter, Israt Jahan, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshiyuki Murata

Bioscience, Biotechnology, and Biochemistry 86 ( 10 ) 1362 - 1367 2022.9

Authorship：Corresponding author Publishing type：Research paper (scientific journal) Publisher：Oxford University Press (OUP)

ABSTRACT

A primary metabolite malate is secreted from guard cells in response to the phytohormone abscisic acid (ABA) and elevated CO2. The secreted malate subsequently facilitates stomatal closure in plants. Here, we investigated the molecular mechanism of malate-induced stomatal closure using inhibitors and ABA signaling component mutants of Arabidopsis thaliana. Malate-induced stomatal closure was impaired by a protein kinase inhibitor, K252a, and also by the disruption of a receptor-like kinase GHR1, which mediates activation of calcium ion (Ca2+) channel by reactive oxygen species (ROS) in guard cells. Malate induced ROS production in guard cells while the malate-induced stomatal closure was impaired by a peroxidase inhibitor, salicylhydroxamic acid, but not by the disruption of Nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) oxidases, RBOHD and RBOHF. The malate-induced stomatal closure was impaired by Ca2+ channel blockers, verapamil, and niflumic acid. These results demonstrate that the malate signaling is mediated by GHR1 and ROS in Arabidopsis guard cells.

DOI： 10.1093/bbb/zbac122

Other Link： https://academic.oup.com/bbb/article-pdf/86/10/1362/45978292/zbac122.pdf

Negative regulation of salicylic acid-induced stomatal closure by glutathione in Arabidopsis thaliana Reviewed International coauthorship International journal

Fahmida Akter, Shintaro Munemasa, Toshiyuki Nakamura, Yoshimasa Nakamura, Yoshiyuki Murata

Bioscience, Biotechnology, and Biochemistry 86 ( 10 ) 1378 - 1382 2022.9

Publishing type：Research paper (scientific journal) Publisher：Oxford University Press (OUP)

ABSTRACT

Salicylic acid (SA) is a ubiquitous phenolic phytohormone that induces stomatal closure. Glutathione (GSH) negatively regulates stomatal closure induced by other plant hormones such as abscisic acid (ABA) and methyl jasmonate (MeJA). However, the involvement of GSH in SA-induced stomatal closure is still unknown. We investigated the regulation of SA signaling by GSH in guard cells using an Arabidopsis thaliana mutant, cad2-1, which is deficient in the first GSH biosynthesis enzyme, γ-glutamylcysteine synthetase. Application of SA decreased stomatal apertures with decreasing intracellular GSH level in guard cells. Decreasing GSH by the cad2-1 mutation and by a GSH-decreasing chemical, 1-chloro-2,4-dinitrobenzene, enhanced the SA-induced stomatal closure. Treatment with glutathione monoethyl ester restored the GSH level in the cad2-1 guard cells and complemented the stomatal phenotype of the mutant. These results indicate that GSH negatively modulates SA-induced stomatal closure in A. thaliana.

DOI： 10.1093/bbb/zbac116

Other Link： https://academic.oup.com/bbb/article-pdf/86/10/1378/45978264/zbac116.pdf

Benzyl isothiocyanate and its metabolites inhibit cell proliferation through protein modification in mouse preosteoclast RAW264.7 cells. Reviewed International coauthorship International journal

Toshiyuki Nakamura, Chiharu Tsutsui, Yu Okuda, Naomi Abe-Kanoh, Saori Okazawa, Shintaro Munemasa, Yoshiyuki Murata, Yoji Kato, Yoshimasa Nakamura

Journal of biochemical and molecular toxicology e23184 2022.8

Language：English Publishing type：Research paper (scientific journal)

Benzyl isothiocyanate (BITC), derived from cruciferous vegetables, is an organosulfur compound exerting antiproliferative effects in several human cancer cells. In this study, we assessed BITC as a potential osteoclastogenesis inhibitor and investigated its underlying mechanism. BITC at 5 μM significantly decreased the viability of the osteoclast-like differentiating RAW264.7 cells, coinciding with the downregulation of the primary biomarkers for osteoclast differentiation, such as the tartrate-resistant acid phosphatase activity and nuclear factor of activated T-cells gene expression. Not only BITC but also its metabolites, inhibited cell proliferation in the normal RAW264.7 cells, suggesting that BITC shows an anti-osteoclastogenesis effect in vivo after its ingestion and metabolism, possibly through an antiproliferative action. Both BITC and its metabolites also enhanced the DNA fragmentation and the caspase-3 activity, whereas their higher concentrations tended to suppress these effects. BITC was intracellularly accumulated when the cells were treated with its metabolites via their degradation into the free form. A quantitative experiment using the proteolysis/high performance liquid chromatography technique showed that the amount of BITC-lysine thiourea in the cells was also increased in a time-dependent manner, suggesting that lysine modification of the cellular proteins actually took place in the cells treated by BITC. Among the cellular proteins, the cleaved caspase-3 was identified as a potential target for lysine modification by BITC. Taken together, BITC dissociated from its metabolites as well as its free form might modulate osteoclastogenesis, possibly through inhibition of cell proliferation by protein modification.

DOI： 10.1002/jbt.23184

Green Tea Catechins, (-)-Catechin Gallate, and (-)-Gallocatechin Gallate are Potent Inhibitors of ABA-Induced Stomatal Closure. Reviewed International coauthorship International journal

Kanane Sato, Shunya Saito, Kohsuke Endo, Masaru Kono, Taishin Kakei, Haruka Taketa, Megumi Kato, Shin Hamamoto, Matteo Grenzi, Alex Costa, Shintaro Munemasa, Yoshiyuki Murata, Yasuhiro Ishimaru, Nobuyuki Uozumi

Advanced science 9 ( 21 ) e2201403 2022.5

Language：English Publishing type：Research paper (scientific journal)

Stomatal movement is indispensable for plant growth and survival in response to environmental stimuli. Cytosolic Ca2+ elevation plays a crucial role in ABA-induced stomatal closure during drought stress; however, to what extent the Ca2+ movement across the plasma membrane from the apoplast to the cytosol contributes to this process still needs clarification. Here the authors identify (-)-catechin gallate (CG) and (-)-gallocatechin gallate (GCG), components of green tea, as inhibitors of voltage-dependent K+ channels which regulate K+ fluxes in Arabidopsis thaliana guard cells. In Arabidopsis guard cells CG/GCG prevent ABA-induced: i) membrane depolarization; ii) activation of Ca2+ permeable cation (ICa ) channels; and iii) cytosolic Ca2+ transients. In whole Arabidopsis plants co-treatment with CG/GCG and ABA suppressed ABA-induced stomatal closure and surface temperature increase. Similar to ABA, CG/GCG inhibited stomatal closure is elicited by the elicitor peptide, flg22 but has no impact on dark-induced stomatal closure or light- and fusicoccin-induced stomatal opening, suggesting that the inhibitory effect of CG/GCG is associated with Ca2+ -related signaling pathways. This study further supports the crucial role of ICa channels of the plasma membrane in ABA-induced stomatal closure. Moreover, CG and GCG represent a new tool for the study of abiotic or biotic stress-induced signal transduction pathways.

DOI： 10.1002/advs.202201403

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