Faculty Profiles - KATSUHARA Maki

Papers - KATSUHARA Maki

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Influence of Low Air Humidity and Low Root Temperature on Water Uptake, Growth and Aquaporin Expression in Rice Plants

Tsuneo Kuwagata, Junko Ishikawa-Sakurai, Hidehiro Hayashi, Kiyoshi Nagasuga, Keiko Fukushi, Arifa Ahamed, Katsuko Takasugi, Maki Katsuhara, Mari Murai-Hatano

PLANT AND CELL PHYSIOLOGY 53 ( 8 ) 1418 - 1431 2012.8

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

The effects of low air humidity and low root temperature (LRT) on water uptake, growth and aquaporin gene expression were investigated in rice plants. The daily transpiration of the plants grown at low humidity was 1.5- to 2-fold higher than that at high humidity. LRT at 13 degrees C reduced transpiration, and the extent was larger at lower humidity. LRT also reduced total dry matter production and leaf area expansion, and the extent was again larger at lower humidity. These observations suggest that the suppression of plant growth by LRT is associated with water stress due to decreased water uptake ability of the root. On the other hand, the net assimilation rate was not affected by low humidity and LRT, and water use efficiency was larger for LRT. We found that low humidity induced coordinated up-regulation of many PIP and TIP aquaporin genes in both the leaves and the roots. Expression levels of two root-specific aquaporin genes, OsPIP2;4 and OsPIP2;5, were increased significantly after 6 and 13 d of LRT exposure. Taken together, we discuss the possibility that aquaporins are part of an integrated response of this crop to low air humidity and LRT.

DOI： 10.1093/pcp/pcs087

Hydrogen peroxide permeability of plasma membrane aquaporins of Arabidopsis thaliana

Cortwa Hooijmaijers, Ji Ye Rhee, Kyung Jin Kwak, Gap Chae Chung, Tomoaki Horie, Maki Katsuhara, Hunseung Kang

JOURNAL OF PLANT RESEARCH 125 ( 1 ) 147 - 153 2012.1

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER TOKYO

Although aquaporins have been known to transport hydrogen peroxide (H(2)O(2)) across cell membranes, the H(2)O(2)-regulated expression patterns and the permeability of every family member of the plasma membrane intrinsic protein (PIP) toward H(2)O(2) have not been determined. This study investigates the H(2)O(2)-regulated expression levels of all plasma membrane aquaporins of Arabidopsis thaliana (AtPIPs), and determines the permeability of every AtPIP for H(2)O(2) in yeast. Hydrogen peroxide treatment of Arabidopsis down-regulated the expression of AtPIP2 subfamily in roots but not in leaves, whereas the expression of AtPIP1 subfamily was not affected by H(2)O(2) treatment. The growth and survival of yeast cells that expressed AtPIP2;2, AtPIP2;4, AtPIP2;5, or AtPIP2;7 was reduced in the presence of H(2)O(2), while the growth of yeast cells expressing any other AtPIP family member was not affected by H(2)O(2). These results show that only certain isoforms of AtPIPs whose expression is regulated by H(2)O(2) treatment are permeable for H(2)O(2) in yeast cells, and suggest that the integrated regulation of aquaporin expression by H(2)O(2) and the capacity of individual aquaporin to transport H(2)O(2) are important for plant response to H(2)O(2).

DOI： 10.1007/s10265-011-0413-2

Functional characterization of a novel plasma membrane intrinsic protein2 in barley

Mineo Shibasaka, Sizuka Sasano, Sigeko Utsugi, Maki Katsuhara

Plant Signaling and Behavior 7 ( 12 ) 1642 - 1646 2012

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

Water homeostasis is crucial to the growth and survival of plants. Plasma membrane intrinsic proteins (PIPs) have been shown to be primary channels mediating water uptake in plant cells. We characterized a novel PIP2 gene, HvPIP2
8 in barley (Hordeum vulgare). HvPIP2
8 shared 72-76% identity with other HvPIP2s and 74% identity with rice OsPIP2
8. The gene was expressed in all organs including the shoots, roots and pistil at a similar level. When HvPIP2
8 was transiently expressed in onion epidermal cells, it was localized to the plasma membrane. HvPIP2
8 showed transport activity for water in Xenopus oocytes, however its interaction with HvPIP1
2 was not observed. These results suggest that HvPIP2
8 plays a role in water homeostasis although further functional analysis is required in future. © 2012 Landes Bioscience.

DOI： 10.4161/psb.22294

Effect of nutrient deficiencies on the water transport properties in figleaf gourd plants

Ji Ye Rhee, Gap Chae Chung, Maki Katsuhara, Sung-Ju Ahn

HORTICULTURE ENVIRONMENT AND BIOTECHNOLOGY 52 ( 6 ) 629 - 634 2011.12

Language：English Publishing type：Research paper (scientific journal) Publisher：KOREAN SOC HORTICULTURAL SCIENCE

Effects of nitrogen, phosphorus, and potassium deficiencies on water transport properties in figleaf gourd plants were studied. Plants were treated for different period of deficiency and physiological parameters such as stomatal conductance, photosynthesis and transpiration were measured. Cell and root pressure probes were utilized to measure turgor and root pressures, half-times of water exchange and hydraulic conductivities to analyze water transport properties. When plants were grown in nitrogen or phosphorus deficient nutrient solutions, they became insensitive to mercury, suggesting that aquaporin was closed resulting in reduced hydraulic conductivity. Inclusion of tungstate, however, restored the sensitivity of cells to mercury, indicating the importance of internal nutrient concentration, not the incoming nutrient supply. The hydrostatic hydraulic conductivity of roots grown in nitrogen deficient solution, representing apoplastic pathway of water transport, was reduced but this reduction was dramatically recovered by the application of tungstate, indicating the importance of nutrient availability from storage pools in relation to water status of plants.

DOI： 10.1007/s13580-011-0046-3

K+ Transport by the OsHKT2;4 Transporter from Rice with Atypical Na+ Transport Properties and Competition in Permeation of K+ over Mg2+ and Ca2+ Ions

Tomoaki Horie, Dennis E. Brodsky, Alex Costa, Toshiyuki Kaneko, Fiorella Lo Schiavo, Maki Katsuhara, Julian I. Schroeder

PLANT PHYSIOLOGY 156 ( 3 ) 1493 - 1507 2011.7

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

Members of class II of the HKT transporters, which have thus far only been isolated from grasses, were found to mediate Na+-K+ cotransport and at high Na+ concentrations preferred Na+-selective transport, depending on the ionic conditions. But the physiological functions of this K+-transporting class II of HKT transporters remain unknown in plants, with the exception of the unique class II Na+ transporter OsHKT2;1. The genetically tractable rice (Oryza sativa; background Nipponbare) possesses two predicted K+-transporting class II HKT transporter genes, OsHKT2;3 and OsHKT2;4. In this study, we have characterized the ion selectivity of the class II rice HKT transporter OsHKT2;4 in yeast and Xenopus laevis oocytes. OsHKT2;4 rescued the growth defect of a K+ uptake-deficient yeast mutant. Green fluorescent protein-OsHKT2;4 is targeted to the plasma membrane in transgenic plant cells. OsHKT2;4-expressing oocytes exhibited strong K+ permeability. Interestingly, however, K+ influx in OsHKT2;4-expressing oocytes did not require stimulation by extracellular Na+, in contrast to other class II HKT transporters. Furthermore, OsHKT2;4-mediated currents exhibited permeabilities to both Mg2+ and Ca2+ in the absence of competing K+ ions. Comparative analyses of Ca2+ and Mg2+ permeabilities in several HKT transporters, including Arabidopsis thaliana HKT1;1 (AtHKT1;1), Triticum aestivum HKT2;1 (TaHKT2;1), OsHKT2;1, OsHKT2;2, and OsHKT2;4, revealed that only OsHKT2;4 and to a lesser degree TaHKT2;1 mediate Mg2+ transport. Interestingly, cation competition analyses demonstrate that the selectivity of both of these class II HKT transporters for K+ is dominant over divalent cations, suggesting that Mg2+ and Ca2+ transport via OsHKT2;4 may be small and would depend on competing K+ concentrations in plants.

DOI： 10.1104/pp.110.168047

Mechanisms of Water Transport Mediated by PIP Aquaporins and Their Regulation Via Phosphorylation Events Under Salinity Stress in Barley Roots

Tomoaki Horie, Toshiyuki Kaneko, Genki Sugimoto, Shizuka Sasano, Sanjib Kumar Panda, Mineo Shibasaka, Maki Katsuhara

PLANT AND CELL PHYSIOLOGY 52 ( 4 ) 663 - 675 2011.4

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

Water homeostasis is crucial to the growth and survival of plants under water-related stress. Plasma membrane intrinsic proteins (PIPs) have been shown to be primary channels mediating water uptake in plant cells. Here we report the water transport activity and mechanisms for the regulation of barley (Hordeum vulgare) PIP aquaporins. HvPIP2 but not HvPIP1 channels were found to show robust water transport activity when expressed alone in Xenopus laevis oocytes. However, the co-expression of HvPIP1 with HvPIP2 in oocytes resulted in significant increases in activity compared with the expression of HvPIP2 alone, suggesting the participation of HvPIP1 in water transport together with HvPIP2 presumably through heteromerization. Severe salinity stress (200 mM NaCl) significantly reduced root hydraulic conductivity (Lp(r)) and the accumulation of six of 10 HvPIP mRNAs. However, under relatively mild stress (100 mM NaCl), only a moderate reduction in Lp(r) with no significant difference in HvPIP mRNA levels was observed. Sorbitol-mediated osmotic stress equivalent to 100 and 200 mM NaCl induced nearly identical Lp(r) reductions in barley roots. Furthermore, the water transport activity in intact barley roots was suggested to require phosphorylation that is sensitive to a kinase inhibitor, staurosporine. HvPIP2s also showed water efflux activity in Xenopus oocytes, suggesting a potential ability to mediate water loss from cells under hypertonic conditions. Water transport via HvPIP aquaporins and the significance of reductions of Lp(r) in barley plants during salinity stress are discussed.

DOI： 10.1093/pcp/pcr027

Rice sodium-insensitive potassium transporter, OsHAK5, confers increased salt tolerance in tobacco BY2 cells

Tomoaki Horie, Mitsuo Sugawara, Tomoyuki Okada, Koichiro Taira, Pulla Kaothien-Nakayama, Maki Katsuhara, Atsuhiko Shinmyo, Hideki Nakayama

JOURNAL OF BIOSCIENCE AND BIOENGINEERING 111 ( 3 ) 346 - 356 2011.3

Language：English Publishing type：Research paper (scientific journal) Publisher：SOC BIOSCIENCE BIOENGINEERING JAPAN

Potassium ion (K(+)) plays vital roles in many aspects of cellular homeostasis including competing with sodium ion (Na(+)) during potassium starvation and salt stress. Therefore, one way to engineer plant cells with improved salt tolerance is to enhance K(+) uptake activity of the cells, while keeping Na(+) out during salt stress. Here, in search for Na(+)-insensitive transporter for this purpose, bacterial expression system was used to characterize two K(+) transporters, OsHAK2 and OsHAK5, isolated from rice (Oryza sativa cv. Nipponbare). The two OsHAK transporters are members of a KT/HAK/KUP transporter family, which is one of the major K(+) transporter families in bacteria, fungi and plants. When expressed in an Escherichia cola K(+) transport mutant strain LB2003, both OsHAK transporters rescued the growth defect in K(+)-limiting conditions by significantly increasing the K(+) content of the cells. Under the condition with a large amount of extracellular Na(+), we found that OsHAK5 functions as a Na(+)-insensitive K(+) transporter, while OsHAK2 is sensitive to extracellular Na(+) and exhibits higher Na(+) over K(+) transport activities. Moreover, constitutive expression of OsHAK5 in cultured-tobacco BY2 (Nicotiana tabacum cv. Bright Yellow 2) cells enhanced the accumulation of K(+) but not Na(+) in the cells during salt stress and conferred increased salt tolerance to the cells. Transient expression experiment indicated that OsHAK5 is localized to the plant plasma membrane. These results suggest that the plasma-membrane localized Na(+) insensitive K(+) transporters, similar to OsHAK5 identified here, could be used as a tool to enhance salt tolerance in plant cells. (C) 2010, The Society for Biotechnology, japan. All rights reserved.

DOI： 10.1016/j.jbiosc.2010.10.014

Hormonal treatment of the bark of rubber trees (Hevea brasiliensis) increases latex yield through latex dilution in relation with the differential expression of two aquaporin genes

Kessarin Tungngoen, Unchera Viboonjun, Panida Kongsawadworakul, Maki Katsuhara, Jean-Louis Julien, Soulaiman Sakr, Herve Chrestin, Jarunya Narangajavana

JOURNAL OF PLANT PHYSIOLOGY 168 ( 3 ) 253 - 262 2011.2

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

Natural rubber is synthesized in laticifers in the inner liber of the rubber tree (Hevea brasiliensis). Upon bark tapping, the latex is expelled due to fiber turgor pressure. The mature laticifers are devoid of plasmodesmata; therefore a corresponding decrease-in the total latex solid content is likely to occur due to water influx inside the laticifers. Auxins and ethylene used as efficient yield stimulants in mature untapped rubber trees, but, bark treatments with abscisic acid (ABA) and salicylic acid (SA) could also induce a transient increase latex yield. We recently reported that there are three aquaporin genes, HbPIP2;1, HbT1P1;1 and HbPIP1;1, that are regulated differentially after ethylene bark treatment. HbPIP2:1 was up-regulated in both the laticifers and the inner liber tissues, whereas HbT1P1:1 was up-regulated in the latex cells, but very markedly down-regulated in the inner liber tissues. Conversely, HbPIP1:1 was down-regulated in both tissues. In the present study, HbPIP2;1 and HbT1P1:1 showed a similar expression in response to auxin, ABA and SA, as seen in ethylene stimulation, while HbPIP1;1 was slightly regulated by auxin, but neither by ABA nor SA. The analysis of the HbPIP1:1 promoter region indicated the presence of only ethylene and auxin responsive elements. In addition, the poor efficiency of this HbPIP1;1 in increasing plasmalemma water conductance was confirmed in Xenopus oocytes. Thus, an increase in latex yield in response to all of these hormones was proposed to be the major function of aquaporins, HbPIP2:1 and HbTIP1;1. This study emphasized that the circulation of water between the laticifers and their surrounding tissues that result in latex dilution, as well as the probable maintenance of the liber tissues turgor pressure, favor the prolongation of latex flow. (C) 2010 Elsevier GmbH. All rights reserved.

DOI： 10.1016/j.jplph.2010.06.009

Abiotic stresses modulate expression of major intrinsic proteins in barley (Hordeum vulgare)

Ayalew Ligaba, Maki Katsuhara, Mineo Shibasaka, Gemechis Djira

COMPTES RENDUS BIOLOGIES 334 ( 2 ) 127 - 139 2011.2

Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER FRANCE-EDITIONS SCIENTIFIQUES MEDICALES ELSEVIER

In one of the most important crops, barley (Hordeum vulgare L), gene expression and physiological roles of most major intrinsic proteins (MIPs) remained to be elucidated. Here we studied expression of five tonoplast intrinsic protein isoforms (HvTIP1;2, HvTIP2:1, HvTIP2;2,HvTIP2;3 and HvTIP4:1), a NOD26-like intrinsic protein (HvNIP2;1) and a plasma membrane intrinsic protein (HvPIP2;1) by using the quantitative real-time RT-PCR. Five-day-old seedlings were exposed to abiotic stresses (salt, heavy metals and nutrient deficiency), abscisic acid (ABA) and gibberellic acid (GA) for 24 h. Treatment with 100 mM NaCl, 0.1 mM ABA and 1 mM GA differentially regulated gene expression in roots and shoots. Nitrogen and prolonged P-deficiency downregulated expression of most MIP genes in roots. Intriguingly, gene expression was restored to the values in the control three days after nutrient supply was resumed. Heavy metals (0.2 mM each of Cd, Cu, Zn and Cr) downregulated the transcript levels by 60-80% in roots, whereas 0.2 mM Hg upregulated expressions of most genes in roots. This was accompanied by a 45% decrease in the rate of transpiration. In order to study the physiological role of the MIPs, cDNA of three genes (HvTIP2:1, HvTIP2:3 and HvNIP2;1) have been cloned and heterologous expression was performed in Xenopus laevis oocytes. Osmotic water permeability was determined by a swelling assay. However, no water uptake activity was observed for the three proteins. Hence, the possible physiological role of the proteins is discussed. (C) 2010 Academie des sciences. Published by Elsevier Masson SAS. All rights reserved.

DOI： 10.1016/j.crvi.2010.11.005

Early response in water relations influenced by NaCl reflects tolerance or sensitivity of barley plants to salinity stress via aquaporins

Maki Katsuhara, Ji Ye Rhee, Genki Sugimoto, Gap Chae Chung

SOIL SCIENCE AND PLANT NUTRITION 57 ( 1 ) 50 - 60 2011.2

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

Barley varieties, K305 and I743, which are sodium chloride (NaCl) tolerant and sensitive respectively, were hydroponically grown to determine the short-term effects of NaCl on the cell water relations in roots using a cell pressure probe, and on the transcript levels of 10 barley PIP aquaporin genes (HvPIPs) in roots. Stomatal conductance, as an indicator of sensitivity to NaCl, was decreased to less than half values of control upon exposure to 100 mmol L-1 NaCl for 24 h in I743 whereas tolerant variety, K305, was able to maintain original conductance. Osmotic half-times of water exchange in cortical cells allowed for a clear distinction between the two varieties up to 200 mmol L-1 NaCl. With treatment duration of up to 12 h with 100 mmol L-1 NaCl, the elastic modulus was reduced in I743 but increased in K305. Hydrostatic half-times of water exchange in K305 increased rapidly, whereas this value remained unchanged in I743. Application of abscisic acid (ABA) after 1 h NaCl treatment restored the hydraulic conductivity of cells (Lp) in K305 but not in I743 whereas the opposite results were obtained when mercury chloride (HgCl2) was applied, verifying the contrasting gating response of aquaporins in two varieties. Reduced expression of HvPIPs was consistent with the reduction of hydraulic conductivity of both varieties after 24 h NaCl, but without any significant differences between them, indicating the importance of the activities of existing aquaporins rather than de novo synthesis to cope with short-term effects of salt stress.

DOI： 10.1080/00380768.2010.541870

Water and H2O2 Permeability of Aquaporins Isolated from Cucumber and Fig leaf Gourd

Ji Ye Rhee, Maki Katsuhara, Hunseung Kang, Gap Chae Chung

HORTICULTURE ENVIRONMENT AND BIOTECHNOLOGY 51 ( 3 ) 167 - 172 2010.6

Language：English Publishing type：Research paper (scientific journal) Publisher：KOREAN SOC HORTICULTURAL SCIENCE

Cucumber aquaporin gene, CsPIP1;1 and figleaf gourd aquaporin gene CfPIP2;1 were injected to Xenopus oocytes in order to study water transport activity. While CsPIP1;1 did not transport any water and was not responsive to mercury, oocyte injected with CfPIP2;1 rapidly increased their volume without saturation indicating active water transport. Calculation of water permeability of oocytes clearly distinguished the two genes. In response to exogenous application of H2O2, only CfPIP2;1 responded, reducing osmotic water permeability. Transgenic Arabidopsis plants overexpressing aquaporin genes did not respond to applied H2O2.

Insights into the salt tolerance mechanism in barley (Hordeum vulgare) from comparisons of cultivars that differ in salt sensitivity

Ayalew Ligaba, Maki Katsuhara

JOURNAL OF PLANT RESEARCH 123 ( 1 ) 105 - 118 2010.1

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER TOKYO

Although barley (Hordeum vulgare L.) is a salt-tolerant crop, the underlying physiological and molecular mechanisms of salt tolerance remain to be elucidated. Therefore, we investigated the response of salt-tolerant (K305) and salt-sensitive (I743) cultivars to salt stress at both physiological and molecular levels. Salt treatment increased xylem sap osmolarity, which was attributed primarily to a rise in Na(+) and Cl(-) concentration; enhanced accumulation of the ions in shoots; and reduced plant growth more severely in I743 than K305. The concentration of K(+) in roots and shoots decreased during 8 h of salt treatment in both cultivars but with no marked difference between cultivars. Hence, the severe growth reduction in I743 is attributed to the elevated levels of (mainly) Na(+) in shoots. Analysis of gene expression using quantitative RT-PCR showed that transcripts of K(+)-transporters (HvHAK1 and HvAKT1), vacuolar H(+)-ATPase and inorganic pyrophosphatase (HvHVA/68 and HvHVP1) were more abundant in shoots of K305 than in shoots of I743. Expression of HvHAK1 and Na(+)/H(+) antiporters (HvNHX1, HvNHX3 and HvNHX4) was higher in roots of K305 than in I743 with prolonged exposure to salt. Taken together, these results suggest that the better performance of K305 compared to I743 during salt stress may be related to its greater ability to sequester Na(+) into sub-cellular compartments and/or maintain K(+) homeostasis.

DOI： 10.1007/s10265-009-0272-2

Differential Sodium and Potassium Transport Selectivities of the Rice OsHKT2;1 and OsHKT2;2 Transporters in Plant Cells

Xuan Yao, Tomoaki Horie, Shaowu Xue, Ho-Yin Leung, Maki Katsuhara, Dennis E. Brodsky, Yan Wu, Julian I. Schroeder

PLANT PHYSIOLOGY 152 ( 1 ) 341 - 355 2010.1

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

Na+ and K+ homeostasis are crucial for plant growth and development. Two HKT transporter/channel classes have been characterized that mediate either Na+ transport or Na+ and K+ transport when expressed in Xenopus laevis oocytes and yeast. However, the Na+/K+ selectivities of the K+-permeable HKT transporters have not yet been studied in plant cells. One study expressing 5' untranslated region-modified HKT constructs in yeast has questioned the relevance of cation selectivities found in heterologous systems for selectivity predictions in plant cells. Therefore, here we analyze two highly homologous rice (Oryza sativa) HKT transporters in plant cells, OsHKT2;1 and OsHKT2;2, that show differential K+ permeabilities in heterologous systems. Upon stable expression in cultured tobacco (Nicotiana tabacum) Bright-Yellow 2 cells, OsHKT2;1 mediated Na+ uptake, but little Rb+ uptake, consistent with earlier studies and new findings presented here in oocytes. In contrast, OsHKT2; 2 mediated Na+-K+ cotransport in plant cells such that extracellular K+ stimulated OsHKT2;2-mediated Na+ influx and vice versa. Furthermore, at millimolar Na+ concentrations, OsHKT2; 2 mediated Na+ influx into plant cells without adding extracellular K+. This study shows that the Na+/K+ selectivities of these HKT transporters in plant cells coincide closely with the selectivities in oocytes and yeast. In addition, the presence of external K+ and Ca2+ down-regulated OsHKT2;1-mediated Na+ influx in two plant systems, Bright-Yellow 2 cells and intact rice roots, and also in Xenopus oocytes. Moreover, OsHKT transporter selectivities in plant cells are shown to depend on the imposed cationic conditions, supporting the model that HKT transporters are multi-ion pores.

DOI： 10.1104/pp.109.145722

A bacterial biosensor for oxidative stress using the constitutively expressed redox-sensitive protein roGFP2. Reviewed International journal

Arias-Barreiro CR, Okazaki K, Koutsaftis A, Inayat-Hussain SH, Tani A, Katsuhara M, Kimbara K, Mori IC

Sensors (Basel, Switzerland) 10 ( 7 ) 6290 - 6306 2010

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

A highly specific, high throughput-amenable bacterial biosensor for chemically induced cellular oxidation was developed using constitutively expressed redox-sensitive green fluorescent protein roGFP2 in E. coli (E. coli-roGFP2). Disulfide formation between two key cysteine residues of roGFP2 was assessed using a double-wavelength ratiometric approach. This study demonstrates that only a few minutes were required to detect oxidation using E. coli-roGFP2, in contrast to conventional bacterial oxidative stress sensors. Cellular oxidation induced by hydrogen peroxide, menadione, sodium selenite, zinc pyrithione, triphenyltin and naphthalene became detectable after 10 seconds and reached the maxima between 80 to 210 seconds, contrary to Cd(2+), Cu(2+), Pb(2+), Zn(2+) and sodium arsenite, which induced the oxidation maximum immediately. The lowest observable effect concentrations (in ppm) were determined as 1.0 × 10(-7) (arsenite), 1.0 × 10(-4) (naphthalene), 1.0 × 10(-4) (Cu(2+)), 3.8 × 10(-4) (H(2)O(2)), 1.0 × 10(-3) (Cd(2+)), 1.0 × 10(-3) (Zn(2+)), 1.0 × 10(-2) (menadione), 1.0 (triphenyltin), 1.56 (zinc pyrithione), 3.1 (selenite) and 6.3 (Pb(2+)), respectively. Heavy metal-induced oxidation showed unclear response patterns, whereas concentration-dependent sigmoid curves were observed for other compounds. In vivo GSH content and in vitro roGFP2 oxidation assays together with E. coli-roGFP2 results suggest that roGFP2 is sensitive to redox potential change and thiol modification induced by environmental stressors. Based on redox-sensitive technology, E. coli-roGFP2 provides a fast comprehensive detection system for toxicants that induce cellular oxidation.

DOI： 10.3390/s100706290

Regulation mechanism of water permeability under hypertonic stress in barley root

Kaneko Toshiyuki, Horie Tomoaki, Shibasaka Mineo, Katsuhara Maki

Plant and Cell Physiology Supplement 2010 0034 - 0034 2010

Publisher：The Japanese Society of Plant Physiologists

The salt stress is one of the representative abiotic stresses that decreases growth and productivity of crops plants. The root water permeability of three barley (Hordeum Vulgare) cultivars that show a difference in the salt sensitivity was measured under salinity stress using a pressure chamber. Water permeability of salt sensitive variety did not show any difference in response to salt stress, in contrast to significant reductions of water permeability found in more salt tolerant varieties. Root water permeability of tolerant cultivars exposed to 100 mM NaCl was found to be extremely down-regulated after 1h. Interestingly, however, water permeability was partially recovered after 4 h, which was followed by a re-down-regulation afterwards. Further measurements using various inhibitors indicated that at least protein phosphorylation and recycling plays an essential role in the regulation mechanism.

DOI： 10.14841/jspp.2010.0.0034.0

Involvement of HbPIP2;1 and HbTIP1;1 Aquaporins in Ethylene Stimulation of Latex Yield through Regulation of Water Exchanges between Inner Liber and Latex Cells in Hevea brasiliensis

Kessarin Tungngoen, Panida Kongsawadworakul, Unchera Viboonjun, Maki Katsuhara, Nicole Brunel, Soulaiman Sakr, Jarunya Narangajavana, Herve Chrestin

PLANT PHYSIOLOGY 151 ( 2 ) 843 - 856 2009.10

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

Natural rubber is synthesized in specialized articulated cells (laticifers) located in the inner liber of Hevea brasiliensis. Upon bark tapping, the laticifer cytoplasm (latex) is expelled due to liber tissue turgor pressure. In mature virgin (untapped) trees, short-term kinetic studies confirmed that ethylene, the rubber yield stimulant used worldwide, increased latex yield, with a concomitant decrease in latex total solid content, probably through water influx in the laticifers. As the mature laticifers are devoid of plasmodesmata, the rapid water exchanges with surrounding liber cells probably occur via the aquaporin pathway. Two full-length aquaporin cDNAs (HbPIP2;1 and HbTIP1;1, for plasma membrane intrinsic protein and tonoplast intrinsic protein, respectively) were cloned and characterized. The higher efficiency of HbPIP2;1 than HbTIP1;1 in increasing plasmalemma water conductance was verified in Xenopus laevis oocytes. HbPIP2;1 was insensitive to HgCl(2). In situ hybridization demonstrated that HbPIP2;1 was expressed in all liber tissues in the young stem, including the laticifers. HbPIP2;1 was up-regulated in both liber tissues and laticifers, whereas HbTIP1;1 was down-regulated in liber tissues but up-regulated in laticifers in response to bark Ethrel treatment. Ethylene-induced HbPIP2;1 up-regulation was confirmed by western-blot analysis. The promoter sequences of both genes were cloned and found to harbor, among many others, ethylene-responsive and other chemical-responsive (auxin, copper, and sulfur) elements known to increase latex yield. Increase in latex yield in response to ethylene was emphasized to be linked with water circulation between the laticifers and their surrounding tissues as well as with the probable maintenance of liber tissue turgor, which together favor prolongation of latex flow.

DOI： 10.1104/pp.109.140228

New production method for greening concrete blocks.

Katsuhara M

Environment Solution Technology 8 ( 3 ) 32 - 36 2009

Biomarkers of Green Roof Vegetation: Anthocyanin and Chlorophyll as Stress Marker Pigments for Plant Stresses of Rooftop Environments.

Mori, I.C, Utsugi,S, Tanakamaru, S, Tani, A, Enomoto, T, Katsuhara, M

Journal of Environmental Engineering and Management 19 21 - 27 2009

Function and regulation of PIP-type water channels in roots of barley under salinity stress

Horie Tomoaki, Kaneko Toshiyuki, Sugimoto Genki, Shibasaka Mineo, Katsuhara Maki

Plant and Cell Physiology Supplement 2009 S0029 - S0029 2009

Publisher：The Japanese Society of Plant Physiologists

The plasma membrane intrinsic protein (PIP) forms the pore for water transport as a water channel in plant cells. We have studied the molecular mechanism of water transport in roots of barley (Hordeum Vulgare cv. Haruna-nijo). Quantitative RT-PCR analyses using roots of barley plants revealed significant reductions in the accumulation of 6 out of 10 HvPIP mRNA in response to 200 mM NaCl. Characterization of water permeability of barely roots using a pressure chamber showed that water permeability was extremely down-regulated in 200 mM NaCl condition. Furthermore, phosphorylation was found to regulate water transport activity. HvPIP1 proteins did not show apparent water transport activity when they were expressed alone in Xenopus laevis oocytes. However, co-expression of HvPIP1 proteins with HvPIP2 proteins was shown to enhance water transport activity of HvPIP2 proteins. The regulation mechanism of water transport via HvPIP proteins in barley roots under salinity stress will be discussed.

DOI： 10.14841/jspp.2009.0.s0029.0

Characterization of four plasma membrane aquaporins in tulip petals: A putative homolog is regulated by phosphorylation

Abul Kalam Azad, Maki Katsuhara, Yoshihiro Sawa, Takahiro Ishikawa, Hitoshi Shibata

PLANT AND CELL PHYSIOLOGY 49 ( 8 ) 1196 - 1208 2008.8

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

We suggested previously that temperature-dependent tulip (Tulipa gesneriana) petal movement that is concomitant with water transport is regulated by reversible phosphorylation of an unidentified plasma membrane intrinsic protein (PIP). In this study, four full-length cDNAs of PIPs from tulip petals were identified and cloned. Two PIPs, namely TgPIP1;1 and TgPIP1;2, are members of the PIP1 subfamily, and the remaining two PIPs, namely TgPIP2;1 and TgPIP2;2, belong to the PIP2 subfamily of aquaporins and were named according to the nomenclature of PIP genes in plants. Of these four homologs, only TgPIP2;2 displayed significant water channel activity in the heterologous expression assay using Xenopus laevis oocytes. The water channel activity of this functional isoform was abolished by mercury and was affected by inhibitors of protein kinase and protein phosphatase. Using a site-directed mutagenesis approach to substitute several serine residues with alanine, and assessing water channel activity using the methylotrophic yeast Pichia pastoris expression assay, we showed that Ser35, Ser116 and Ser274 are the putative phosphorylation sites of TgPIP2;2. Real-time reverse transcriptionPCR analysis revealed that the transcript levels of TgPIP1;1 and TgPIP1;2 in tulip petals, stems, leaves, bulbs and roots are very low when compared with those of TgPIP2;1 and TgPIP2;2. The transcript level of TgPIP2;1 is negligible in roots, and TgPIP2;2 is ubiquitously expressed in all organs with significant transcript levels. From the data reported herein, we suggest that TgPIP2;2 might be modulated by phosphorylation and dephosphorylation for regulating water channel activity, and may play a role in transcellular water transport in all tulip organs.

DOI： 10.1093/pcp/pcn095

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