| kensuke kusumi | Last modified date：2023.12.06 |
Lecturer /
Informational biology /
Department of Biology /
Faculty of Sciences
Papers
| 1. | Kazuhiro Ito, Doshun Ito, Mina Goto, Sae Suzuki, Shinji Masuda, Koh Iba, Kensuke Kusumi, Regulation of ppGpp synthesis and its impact on chloroplast biogenesis during early leaf development in rice, Plant and Cell Physiology, 10.1093/pcp/pcac053, 63, 7, 919-931, 2022.04, [URL], Guanosine tetraphosphate (ppGpp) is known as an alarmone that mediates bacterial stress responses. In plants, ppGpp is synthesized in chloroplasts from GTP and ATP and functions as a regulator of chloroplast gene expression to affect photosynthesis and plant growth. This observation indicates that ppGpp metabolism is closely related to chloroplast function, but the regulation of ppGpp and its role in chloroplast differentiation are not well understood. In rice, ppGpp directly inhibits plastidial guanylate kinase (GKpm), a key enzyme in GTP biosynthesis. GKpm is highly expressed during early leaf development in rice, and the GKpm-deficient mutant, virescent-2 (v2), develops chloroplast-deficient chlorotic leaves under low-temperature conditions. To examine the relationship between GTP synthesis and ppGpp homeostasis, we generated transgenic rice plants over-expressing RSH3, a protein known to act as a ppGpp synthase. When RSH3 was overexpressed in v2, the leaf chlorosis was more severe. Although the RSH3 overexpression in the wild type caused no visible effects, pulse amplitude modulation fluorometer measurements indicated that photosynthetic rates were reduced in this line. This finding implies that the regulation of ppGpp synthesis in rice is involved in the maintenance of the GTP pool required to regulate plastid gene expression during early chloroplast biogenesis. We further investigated changes in the expressions of RelA/SpoT Homolog (RSH) genes encoding ppGpp synthases and hydrolases during the same period. Comparing the expression of these genes with the cellular ppGpp content suggests that the basal ppGpp level is determined by the antagonistic action of multiple RSH enzymatic activities during early leaf development in rice.. |
| 2. | Wataru Yamori, Kensuke Kusumi, Koh Iba, Ichiro Terashima, Increased stomatal conductance induces rapid changes to photosynthetic rate in response to naturally fluctuating light conditions in rice, Plant Cell and Environment, 10.1111/pce.13725, 2020.01, A close correlation between stomatal conductance and the steady-state photosynthetic rate has been observed for diverse plant species under various environmental conditions. However, it remains unclear whether stomatal conductance is a major limiting factor for the photosynthetic rate under naturally fluctuating light conditions. We analysed a SLAC1 knockout rice line to examine the role of stomatal conductance in photosynthetic responses to fluctuating light. SLAC1 encodes a stomatal anion channel that regulates stomatal closure. Long exposures to weak light before treatments with strong light increased the photosynthetic induction time required for plants to reach a steady-state photosynthetic rate and also induced stomatal limitation of photosynthesis by restricting the diffusion of CO<sub>2</sub> into leaves. The slac1 mutant exhibited a significantly higher rate of stomatal opening after an increase in irradiance than wild-type plants, leading to a higher rate of photosynthetic induction. Under natural conditions, in which irradiance levels are highly variable, the stomata of the slac1 mutant remained open to ensure efficient photosynthetic reaction. These observations reveal that stomatal conductance is important for regulating photosynthesis in rice plants in the natural environment with fluctuating light.. |
| 3. | Juntaro Negi, Shintaro Munemasa, Boseok Song, Ryosuke Tadakuma, Mayumi Fujita, Tamar Azoulay-Shemer, Cawas B. Engineer, Kensuke Kusumi, Ikuo Nishida, Julian I. Schroeder, Koh Iba, Eukaryotic lipid metabolic pathway is essential for functional chloroplasts and CO2 and light responses in Arabidopsis guard cells, Proceedings of the National Academy of Sciences of the United States of America, 10.1073/pnas.1810458115, 115, 36, 9038-9043, 2018.09, Stomatal guard cells develop unique chloroplasts in land plant species. However, the developmental mechanisms and function of chloroplasts in guard cells remain unclear. In seed plants, chloroplast membrane lipids are synthesized via two pathways: the prokaryotic and eukaryotic pathways. Here we report the central contribution of endoplasmic reticulum (ER)-derived chloroplast lipids, which are synthesized through the eukaryotic lipid metabolic pathway, in the development of functional guard cell chloroplasts. We gained insight into this pathway by isolating and examining an Arabidopsis mutant, gles1 (green less stomata 1), which had achlorophyllous stomatal guard cells and impaired stomatal responses to CO<sub>2</sub> and light. The GLES1 gene encodes a small glycine-rich protein, which is a putative regulatory component of the trigalactosyldiacylglycerol (TGD) protein complex that mediates ER-to-chloroplast lipid transport via the eukaryotic pathway. Lipidomic analysis revealed that in the wild type, the prokaryotic pathway is dysfunctional, specifically in guard cells, whereas in gles1 guard cells, the eukaryotic pathway is also abrogated. CO<sub>2</sub>-induced stomatal closing and activation of guard cell S-type anion channels that drive stomatal closure were disrupted in gles1 guard cells. In conclusion, the eukaryotic lipid pathway plays an essential role in the development of a sensing/ signaling machinery for CO<sub>2</sub> and light in guard cell chloroplasts.. |
| 4. | Kensuke Kusumi, Ayana Hashimura, Yoshiko Yamamoto, Juntaro Negi, Koh Iba, Contribution of the S-type anion channel SLAC1 to stomatal control and its dependence on developmental stage in rice, Plant and Cell Physiology, 10.1093/pcp/pcx142, 58, 12, 2085-2094, 2017.06, Rice production depends on water availability and carbon fixation by photosynthesis. Therefore, optimal control of stomata, which regulate leaf transpiration and CO2 absorption, is important for high productivity. SLOW ANION CHANNEL-ASSOCIATED 1 (SLAC1) is an S-type anion channel protein that controls stomatal closure in response to elevated CO2. Rice slac1 mutants showed significantly increased stomatal conductance (gs) and enhanced CO2 assimilation. To discern the contribution of stomatal regulation to rice growth, we compared gs in the wild type (WT) and two mutants, slac1 and the dominant-positive mutant SLAC1-F461A, which expresses a point mutation causing an amino acid substitution (F461A) in SLAC1, at different growth stages. Because the side group of F461 is estimated to function as the channel gate, stomata in the SLAC1-F461A mutant are expected to close constitutively. All three lines had maximum gs during the tillering stage, when the gs values were 50% higher in slac1 and 70% lower in SLAC1-F461A, compared with the WT. At the tillering stage, the gs values were highest in the first leaves at the top of the stem and lower in the second and third leaves in all three lines. Both slac1 and SLAC1-F461A retained the ability to change gs in response to the day–night cycle, and showed differences in tillering rate and plant height compared with the WT, and lower grain yield. These observations show that SLAC1 plays a crucial role in regulating stomata in rice at the tillering stage.. |
| 5. | Kensuke Kusumi, Koh Iba, Establishment of the chloroplast genetic system in rice during early leaf development and at low temperatures, Frontiers in Plant Science, 10.3389/fpls.2014.00386, 5, 386, 2014.07, [URL], Chloroplasts are the central nodes of the metabolic network in leaf cells of higher plants, and the conversion of proplastids into chloroplasts is tightly coupled to leaf development. During early leaf development, the structure and function of the chloroplasts differ greatly from those in a mature leaf, suggesting the existence of a stage-specific mechanism regulating chloroplast development during this period. Here, we discuss the identification of the genes affected in low temperature-conditional mutants of rice (Oryza sativa). These genes encode factors involved in chloroplast rRNA regulation (NUS1), and nucleotide metabolism in mitochondria, chloroplasts and cytosol (V2, V3, ST1). These genes are all preferentially expressed in the early leaf developmental stage P4, and depleting them causes altered chloroplast transcription and translation, and ultimately leaf chlorosis. Therefore, it is suggested that regulation of cellular nucleotide pools and nucleotide metabolism is indispensable for chloroplast development under low temperatures at this stage. This review summarizes the current understanding of these factors and discusses their roles in chloroplast biogenesis.. |
| 6. | Yuhta Nomura, Atsushi Izumi, Yoshinori Fukunaga, Kensuke Kusumi, Koh Iba, Seiya Watanabe, Yoichi Nakahira, Andreas P.M. Weber, Akira Nozawa, Yuzuru Tozawa, Diversity in guanosine 3′,5′-bisdiphosphate (ppGpp) sensitivity among guanylate kinases of bacteria and plants, Journal of Biological Chemistry, 10.1074/jbc.M113.534768, 289, 22, 15631-15641, 2014.05, Background: The ppGpp signaling system is operative in plant chloroplasts and bacteria. Results: Chloroplast and cytosolic guanylate kinases (GKs) of plants are sensitive and insensitive to ppGpp, respectively, whereas bacterial GKs show diversity in ppGpp sensitivity. Conclusion: GTP biosynthesis in chloroplasts is controlled by ppGpp. Significance: Identification of the targets of ppGpp should provide insight into biological processes regulated by this nucleotide.. |
| 7. | Yuhta Nomura, Atsushi Izumi, Yoshinori Fukunaga, Kensuke Kusumi, Koh Iba, Seiya Watanabe, Yoichi Nakahira, Andreas P. M. Weber, Akira Nozawa, Yuzuru Tozawa, Diversity in Guanosine 3′,5′-Bisdiphosphate (ppGpp) Sensitivity Among Guanylate Kinases of Bacteria and Plants. , Journal of Biological Chemistry, 10.1074/jbc.M113.534768., 289, 15631-15641, 2014.04, [URL], The guanosine 3′,5′-bisdiphosphate (ppGpp) signaling system is shared by bacteria and plant chloroplasts, but its role in plants has remained unclear. Here we show that guanylate kinase (GK), a key enzyme in guanine nucleotide biosynthesis that catalyzes the conversion of GMP to GDP, is a target of regulation by ppGpp in chloroplasts of rice, pea, and Arabidopsis. Plants have two distinct types of GK that are localized to organelles (GKpm) or to the cytosol (GKc), with both enzymes being essential for growth and development. We found that the activity of rice GKpm in vitro was inhibited by ppGpp with a Ki of 2.8 μM relative to the substrate GMP, whereas the Km of this enzyme for GMP was 73 μM. The IC50 of ppGpp for GKpm was ~10 μM. In contrast, the activity of rice GKc was insensitive to ppGpp, as was that of GK from baker′s yeast, which is also a cytosolic enzyme. These observations suggest that ppGpp plays a pivotal role in the regulation of GTP biosynthesis in chloroplasts through specific inhibition of GKpm activity, with the regulation of GTP biosynthesis in chloroplasts thus being independent of that in the cytosol. We also found that GKs of Escherichia coli and Synechococcus elongatus PCC 7942 are insensitive to ppGpp, in contrast to the ppGpp sensitivity of the Bacillus subtilis enzyme. Our biochemical characterization of GK enzymes has thus revealed a novel target of ppGpp in chloroplasts and has uncovered diversity among bacterial GKs with regard to regulation by ppGpp.. |
| 8. | Mimi Hashimoto-Sugimoto, Juntaro Negi, Kensuke Kusumi, Koh Iba, New Approaches to the Biology of Stomatal Guard Cells, 10.1093/pcp/pct145, 55, 2, 241-250, 2014.02, [URL], CO2 acts as an environmental signal that regulates stomatal movements. High CO2 concentrations reduce stomatal aperture, whereas low concentrations trigger stomatal opening. In contrast to our advanced understanding of light and drought stress responses in guard cells, the molecular mechanisms underlying stomatal CO2 sensing and signaling are largely unknown. Leaf temperature provides a convenient indicator of transpiration, and can be used to detect mutants with altered stomatal control. To identify genes that function in CO2 responses in guard cells, CO2-insensitive mutants were isolated through high-throughput leaf thermal imaging. The isolated mutants are categorized into three groups according to their phenotypes: (i) impaired in stomatal opening under low CO2 concentrations; (ii) impaired in stomatal closing under high CO2 concentrations; and (iii) impaired in stomatal development. Characterization of these mutants has begun to yield insights into the mechanisms of stomatal CO2 responses. In this review, we summarize the current status of the field and discuss future prospects.. |
| 9. | Juntaro Negi, Kosuke Moriwaki, Mineko Konishi, Ryusuke Yokoyama, Toshiaki Nakano, Kensuke Kusumi, Mimi Hashimoto-Sugimoto, Julian I. Schroeder, Kazuhiko Nishitani, Shuichi Yanagisawa, Koh Iba, A dof transcription factor, SCAP1, is essential for the development of functional stomata in arabidopsis, Current Biology, 10.1016/j.cub.2013.02.001, 23, 6, 479-484, 2013.03, Stomata are highly specialized organs that consist of pairs of guard cells and regulate gas and water vapor exchange in plants [1-3]. Although early stages of guard cell differentiation have been described [4-10] and were interpreted in analogy to processes of cell type differentiation in animals [11], the downstream development of functional stomatal guard cells remains poorly understood. We have isolated an Arabidopsis mutant, stomatal carpenter 1 (scap1), that develops irregularly shaped guard cells and lacks the ability to control stomatal aperture, including CO2-induced stomatal closing and light-induced stomatal opening. SCAP1 was identified as a plant-specific Dof-type transcription factor expressed in maturing guard cells, but not in guard mother cells. SCAP1 regulates the expression of genes encoding key elements of stomatal functioning and morphogenesis, such as K+ channel protein, MYB60 transcription factor, and pectin methylesterase. Consequently, ion homeostasis was disturbed in scap1 guard cells, and esterification of extracellular pectins was impaired so that the cell walls lining the pores did not mature normally. We conclude that SCAP1 regulates essential processes of stomatal guard cell maturation and functions as a key transcription factor regulating the final stages of guard cell differentiation.. |
| 10. | Negi, J., Moriwaki, K., Konishi, M., Yokoyama, R., Nakano, T., Kusumi, K., Hashimoto-Sugimoto, M., Schroeder, J.I., Nishitani, K., Yanagisawa, S. and Iba, K. , A Dof transcription factor, SCAP1, is essential for the development of functional stomata in Arabidopsis., Current Biology, 10.1093/pcp/pct145, 55, 241-250, 2013.02, Stomata are highly specialized organs that consist of pairs of guard cells and regulate gas and water vapor exchange in plants [1-3]. Although early stages of guard cell differentiation have been described [4-10] and were interpreted in analogy to processes of cell type differentiation in animals [11], the downstream development of functional stomatal guard cells remains poorly understood. We have isolated an Arabidopsis mutant, stomatal carpenter 1 (scap1), that develops irregularly shaped guard cells and lacks the ability to control stomatal aperture, including CO2-induced stomatal closing and light-induced stomatal opening. SCAP1 was identified as a plant-specific Dof-type transcription factor expressed in maturing guard cells, but not in guard mother cells. SCAP1 regulates the expression of genes encoding key elements of stomatal functioning and morphogenesis, such as K(+) channel protein, MYB60 transcription factor, and pectin methylesterase. Consequently, ion homeostasis was disturbed in scap1 guard cells, and esterification of extracellular pectins was impaired so that the cell walls lining the pores did not mature normally. We conclude that SCAP1 regulates essential processes of stomatal guard cell maturation and functions as a key transcription factor regulating the final stages of guard cell differentiation.. |
| 11. | Kusumi, K., Hirotsuka, S., Kumamaru, T. and Iba, K. , Increased leaf photosynthesis caused by elevated stomatal conductance in a rice mutant deficient in SLAC1, a guard cell anion channel protein., Journal of Experimental Botany, 10.1093/jxb/ers216, 63, 15, 5635-5644, 2012.08, [URL], In rice (Oryza sativa L.), leaf photosynthesis is known to be highly correlated with stomatal conductance; however, it remains unclear whether stomatal conductance dominantly limits the photosynthetic rate. SLAC1 is a stomatal anion channel protein controlling stomatal closure in response to environmental [CO2]. In order to examine stomatal limitations to photosynthesis, we isolated and characterized a SLAC1-deficient mutant of rice. A TILLING screen of NMU-derived mutant lines was conducted for the rice SLAC1 ortholog gene Os04g0674700, and four mutant lines containing mutations within the open reading frame were obtained. A second screen using an infrared thermography camera revealed that one of the mutants, named slac1, had a constitutive low-leaf-temperature phenotype. Measurement of leaf gas exchange showed that slac1 plants grown in the greenhouse had significantly higher stomatal conductance (gs), rates of photosynthesis (A) and ratios of internal [CO2] to ambient [CO2] (Ci/Ca) compared to wild-type plants, whereas there was no significant difference in the response of photosynthesis to internal [CO2] (A/Ci curves). These observations demonstrate that in well-watered conditions, stomatal conductance is a major determinant of photosynthetic rate in rice. . |
| 12. | Sakamoto H, Sakata K, Kusumi K, Kojima M, Sakakibara H, Iba K., Interaction between a plasma membrane-localized ankyrin-repeat protein ITN1 and a nuclear protein RTV1., Biochem. Biophys. Res. Commun., 10.1016/j.bbrc.2012.05.136, 423, 2, 392-397, 2012.06, The increased tolerance to NaCl 1 (ITN1) protein is a plasma membrane (PM)-localized protein involved in responses to NaCl stress in Arabidopsis. The predicted structure of ITN1 is composed of multiple transmembrane regions and an ankyrin-repeat domain that is known to mediate protein-protein interactions. To elucidate the molecular functions of ITN1, we searched for interacting partners using a yeast two-hybrid assay, and a nuclear-localized DNA-binding protein, RTV1, was identified as a candidate. Bimolecular fluorescence complementation analysis revealed that RTV1 interacted with ITN1 at the PM and nuclei in vivo. RTV1 tagged with red fluorescent protein localized to nuclei and ITN1 tagged with green fluorescent protein localized to PM; however, both proteins localized to both nuclei and the PM when co-expressed. These findings suggest that RTV1 and ITN1 regulate the subcellular localization of each other.. |
| 13. | Kensuke Kusumi, Chikako Sakata, Takahiro Nakamura, Shinji Kawasaki, Atsushi Yoshimura, Koh Iba, A plastid protein NUS1 is essential for build-up of the genetic system for early chloroplast development under cold stress conditions, Plant Journal, 10.1111/j.1365-313X.2011.04755.x, 68, 6, 1039-1050, 2011.12, During early chloroplast differentiation, the regulation of the plastid genetic system including transcription and translation differs greatly from that in the mature chloroplast, suggesting the existence of a stage-dependent mechanism that regulates the chloroplast genetic system during this period. The virescent-1 (v 1) mutant of rice (Oryza sativa) is temperature-conditional and develops chlorotic leaves under low-temperature conditions. We reported previously that leaf chlorosis in the v 1 mutant is caused by blockage of the activation of the chloroplast genetic system during early leaf development. Here we identify the V 1 gene, which encodes a chloroplast-localized protein NUS1. Accumulation of NUS1 specifically occurred in the pre-emerged immature leaves, and is enhanced by low-temperature treatment. The C-terminus of NUS1 shows structural similarity to the bacterial antitermination factor NusB, which is known to play roles in the regulation of ribosomal RNA transcription. The RNA-immunoprecipitation and gel mobility shift assays indicated that NUS1 binds to several regions of chloroplast RNA including the upstream leader region of the 16S rRNA precursor. In the leaves of the NUS1-deficient mutant, accumulation of chloroplast rRNA during early leaf development was impaired and chloroplast translation/transcription capacity was severely suppressed under low temperature. Our results suggest that NUS1 is involved in the regulation of chloroplast RNA metabolism and promotes the establishment of the plastid genetic system during early chloroplast development under cold stress conditions.. |
| 14. | Kusumi, Kensuke; Sakata, Chikako; Nakamura, Takahiro; Kawasaki, Shinji; Yoshimura, Atsushi; Iba, Koh, A plastid protein NUS1 is essential for build-up of the genetic system for early chloroplast development under cold stress condition, The Plant Journal, 2011.09. |
| 15. | Monda, K., Negi, J., Iio, A., Kusumi, K., Kojima, M., Hashimoto, M., Sakakibara, H. and Iba, K. , Environmental regulation of stomatal response in the Arabidopsis Cvi-0 ecotype. Planta., Planta, 10.1007/s00425-011-1424-x, Online first, 2011.05. |
| 16. | Kusumi, K., Chono, Y., Shimada, H., Gotoh, E., Tsuyama, M. and Iba, K., Chloroplast biogenesis during the early stage of leaf development in rice., Plant Biotechnology, 27, 1, 85-90, 2010.01. |
| 17. | Kusumi, K., Hirotsuka, S., Shimada, H., Chono, Y., Matsuda, O. and Iba, K., Contribution of chloroplast biogenesis to carbon-nitrogen balance during early leaf development in rice, J. Plant Res. , 123, 4, 617-622, 2010.01. |
| 18. | Kensuke Kusumi, Yoko Chono, Hiroshi Shimada, Eiji Gotoh, Michito Tsuyama, Koh Iba, Chloroplast biogenesis during the early stage of leaf development in rice, Plant Biotechnology, 10.5511/plantbiotechnology.27.85, 27, 1, 85-90, 2010.01, In rice, the developmental process in leaf formation can be divided into 7 stages (stages P0 to P6). We investigated chloroplast biogenesis and physiological changes in the developing leaves at the stage P4, during which leaf blade elongation and establishment of basic leaf blade structure occur. Chlorophyll content was negligible in the leaves early in the P4 stage and increased rapidly as they enters the late P4 phase. Chlorophyll fluorescence ratio (Fv/Fm) also increased markedly and the final value was comparable with that of mature leaves. Gene expression analysis showed that during the P4 stage, chloroplasts in the leaf cell undergo all three steps of differentiation: (i) plastid division and DNA replication (ii) establishment of plastid genetic system (iii) activation of photosynthetic apparatus. These observations suggest that the P4 is key in the development of a leaf, during which leaf rapidly differentiated both morphologically and physiologically, and that the P4 leaf is suitable for investigation of physiological relationships between chloroplast and leaf development.. |
| 19. | Kensuke Kusumi, Shoko Hirotsuka, Hiroshi Shimada, Yoko Chono, Osamu Matsuda, Koh Iba, Contribution of chloroplast biogenesis to carbon-nitrogen balance during early leaf development in rice, Journal of Plant Research, 10.1007/s10265-009-0277-x, 123, 4, 617-622, 2010.01, Chloroplast biogenesis is most significant during the changes in cellular organization associated with leaf development in higher plants. To examine the physiological relationship between developing chloroplasts and host leaf cells during early leaf development, we investigated changes in the carbon and nitrogen contents in leaves at the P4 developmental stage of rice, during which leaf blade structure is established and early events of chloroplast differentiation occur. During the P4 stage, carbon content on a dry mass basis remained constant, whereas the nitrogen content decreased by 30%. Among carbohydrates, sucrose and starch accumulated to high levels early in the P4 stage, and glucose, fructose and cellulose degradation increased during the mid-to-late P4 stage. In the chloroplast-deficient leaves of the virescent-1 mutant of rice, however, the carbon and nitrogen contents, as well as the C/N ratio during the P4 stage, were largely unaffected. These observations suggest that developing rice leaves function as sink organs at the P4 stage, and that chloroplast biogenesis and carbon and nitrogen metabolism in the leaf cell is regulated independently at this stage.. |
| 20. | Yoo, S-C., Cho, S-H., Sugimoto, H., Li, Jinjie., Kusumi, K., Koh, H-J., Iba, K. and Paek, N-C. , Rice Virescent-3 and Stripe-1 encoding the large and small subunits of ribonucleotide reductase are required for chloroplast biogenesis during early leaf development., Plant Physiol. , 150: 388-401, 2009.05. |
| 21. | Soo Cheul Yoo, Sung Hwan Cho, Hiroki Sugimoto, Jinjie Li, Kensuke Kusumi, Hee Jong Koh, Koh Iba, Nam Chon Paek, Rice virescent3 and stripe1 encoding the large and small subunits of ribonucleotide reductase are required for chloroplast biogenesis during early leaf development1[w][oa], Plant physiology, 10.1104/pp.109.136648, 150, 1, 388-401, 2009.05, The virescent3 (v3) and stripe1 (st1) mutants in rice (Oryza sativa) produce chlorotic leaves in a growth stage-dependent manner under field conditions. They are temperature-conditional mutants that produce bleached leaves at a constant 20°C or 30°C but almost green leaves under diurnal 30°C/20°C conditions. Here, we show V3 and St1, which encode the large and small subunits of ribonucleotide reductase (RNR), RNRL1, and RNRS1, respectively. RNR regulates the rate of deoxyribonucleotide production for DNA synthesis and repair. RNRL1 and RNRS1 are highly expressed in the shoot base and in young leaves, and the expression of the genes that function in plastid transcription/translation and in photosynthesis is altered in v3 and st1 mutants, indicating that a threshold activity of RNR is required for chloroplast biogenesis in developing leaves. There are additional RNR homologs in rice, RNRL2 and RNRS2, and eukaryotic RNRs comprise α2β 2 heterodimers. In yeast, RNRL1 interacts with RNRS1 (RNRL1:RNRS1) and RNRL2:RNRS2, but no interaction occurs between other combinations of the large and small subunits. The interacting activities are RNRL1:RNRS1 > RNRL1:rnrs1(st1) > rnrl1(v3):RNRS1 > rnrl1(v3):rnrs1(st1), which correlate with the degree of chlorosis for each genotype. This suggests that missense mutations in rnrl1(v3) and rnrs1 (st1) attenuate the first αβ dimerization. Moreover, wild-type plants exposed to a low concentration of an RNR inhibitor, hydroxyurea, produce chlorotic leaves without growth retardation, reminiscent of v3 and st1 mutants. We thus propose that upon insufficient activity of RNR, plastid DNA synthesis is preferentially arrested to allow nuclear genome replication in developing leaves, leading to continuous plant growth.. |
| 22. | Yara, A., Yaeno, T., Hasegawa, M., Seto, H., Seo, S., Kusumi, K. and Iba, K. , Resistance to Magnaporthe grisea in transgenic rice with suppressed expression of genes encoding allene oxide cyclase and phytodienoic acid reductase. , Biochem. Biophys. Res. Commun. , 376: 460-465, 2008.11. |
| 23. | Asanori Yara, Takashi Yaeno, Morifumi Hasegawa, Hideharu Seto, Shigemi Seo, Kensuke Kusumi, Koh Iba, Resistance to Magnaporthe grisea in transgenic rice with suppressed expression of genes encoding allene oxide cyclase and phytodienoic acid reductase, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2008.08.157, 376, 3, 460-465, 2008.11, Linolenic acid (18:3) and its derivative jasmonic acid (JA) are important molecules in disease resistance in many dicotyledonous plants. We have previously used 18:3- and JA-deficient rice (F78Ri) to investigate the roles of fatty acids and their derivatives in resistance to the blast fungus Magnaporthe grisea [A. Yara, T. Yaeno, J.-L. Montillet, M. Hasegawa, S. Seo, K. Kusumi, K. Iba, Enhancement of disease resistance to Magnaporthe grisea in rice by accumulation of hydroxy linoleic acid, Biochem. Biophys. Res. Commun. 370 (2008) 344-347; A. Yara, T. Yaeno, M. Hasegawa, H. Seto, J.-L. Montillet, K. Kusumi, S. Seo, K. Iba, Disease resistance against Magnaporthe grisea is enhanced in transgenic rice with suppression of ω-3 fatty acid desaturases, Plant Cell Physiol. 48 (2007) 1263-1274]. However, because F78Ri plants are suppressed in the first step of the JA biosynthetic pathway, we could not confirm the specific contribution of JA to disease resistance. In this paper, we generated two JA-deficient rice lines (AOCRi and OPRRi) with suppressed expression of the genes encoding allene oxide cyclase (AOC) and 12-oxo-phytodienoic acid reductase (OPR), which catalyze late steps in the JA biosynthetic pathway. The levels of disease resistance in the AOCRi and OPRRi lines were equal to that in wild-type plants. Our data suggest that resistance to M. grisea is not dependent on JA synthesis.. |
| 24. | Yara, A., Yaeno, T., Montillet, J.L., Hasegawa, M., Seo, S., Kusumi, K. and Iba, K. , Enhancement of disease resistance to Magnaporthe grisea in rice by accumulation of hydroxy linoleic acid. , Biochem. Biophys. Res. Commun. , 370: 344-347, 2008.05. |
| 25. | Asanori Yara, Takashi Yaeno, Jean Luc Montillet, Morifumi Hasegawa, Shigemi Seo, Kensuke Kusumi, Koh Iba, Enhancement of disease resistance to Magnaporthe grisea in rice by accumulation of hydroxy linoleic acid, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2008.03.083, 370, 2, 344-347, 2008.05, Linoleic acid (18:2) and linolenic acid (18:3) are sources for various oxidized metabolites called oxylipins, some of which inhibit growth of fungal pathogens. In a previous study, we found disease resistance to rice blast fungus Magnaporthe grisea enhanced in 18:2-accumulating transgenic rice (F78Ri) in which the conversion from 18:2 to 18:3 was suppressed. Here, we demonstrate that 18:2-derived hydroperoxides and hydroxides (HPODEs and HODEs, respectively) inhibit growth of M. grisea more strongly than their 18:3-derived counterparts. Furthermore, in F78Ri plants, the endogenous levels of HPODEs and HODEs increased significantly, compared with wild-type plants. These results suggest that the increased accumulation of antifungal oxylipins, such as HPODEs and HODEs, causes the enhancement of disease resistance against M. grisea.. |
| 26. | Sugimoto, H., Kusumi, K., Noguchi, K., Yano, M., Yoshimura, A., and Iba, K., The rice nuclear gene, VIRESCENT 2, is essential for chloroplast development and encodes a novel type of guanylate kinase targeted to plastids and mitochondria., Plant Journal, 52: 512-527, 2007.11. |
| 27. | Hiroki Sugimoto, Kensuke Kusumi, Ko Noguchi, Masahiro Yano, Atsushi Yoshimura, Koh Iba, The rice nuclear gene, VIRESCENT 2, is essential for chloroplast development and encodes a novel type of guanylate kinase targeted to plastids and mitochondria, Plant Journal, 10.1111/j.1365-313X.2007.03251.x, 52, 3, 512-527, 2007.11, Guanylate kinase (GK) is a critical enzyme in guanine nucleotide metabolism pathways, catalyzing the phosphorylation of (d)GMP to (d)GDP. Here we show that a novel gene, VIRESCENT 2 (V2), encodes a new type of GK (designated pt/mtGK) that is localized in plastids and mitochondria. We initially identified the V2 gene by positional cloning of the rice v2 mutant. The v2 mutant is temperature-sensitive and develops chlorotic leaves at restrictive temperatures. The v2 mutation causes inhibition of chloroplast differentiation; in particular, it disrupts the chloroplast translation machinery during early leaf development [Sugimoto et al. (2004)Plant Cell Physiol. 45, 985]. In the bacterial and animal species studied to date, GK is localized in the cytoplasm and participates in maintenance of the guanine nucleotide pools required for many fundamental cellular processes. Phenotypic analysis of rice seedlings with RNAi knockdown of cytosolic GK (designated cGK) showed that cGK is indispensable for the growth and development of plants, but not for chloroplast development. Thus, rice has two types of GK, as does Arabidopsis, suggesting that higher plants have two types of GK. Our results suggest that, of the two types of GK, only pt/mtGK is essential for chloroplast differentiation.. |
| 28. | Asanori Yara, Takashi Yaeno, Morifumi Hasegawa, Hideharu Seto, Jean Luc Montillet, Kensuke Kusumi, Shigemi Seo, Koh Iba, Disease resistance against Magnaporthe grisea is enhanced in transgenic rice with suppression of ω-3 fatty acid desaturases, Plant and Cell Physiology, 10.1093/pcp/pcm107, 48, 9, 1263-1274, 2007.09, Linolenic acid (18:3) is the most abundant fatty acid in plant membrane lipids and is a source for various oxidized metabolites, called oxylipins. 18:3 and oxylipins play important roles in the induction of defense responses to pathogen infection and wound stress in Arabidopsis. However, in rice, endogenous roles for 18:3 and oxylipins in disease resistance have not been confirmed. We generated 18:3-deficient transgenic rice plants (F78Ri) with co-suppression of two ω-3 fatty acid desaturases, OsFAD7 and OsFAD8. that synthesize 18:3. The F78Ri plants showed enhanced resistance to the phytopathogenic fungus Magnaporthe grisea. A typical 18:3-derived oxylipin, jasmonic acid (JA), acts as a signaling molecule in defense responses to fungal infection in Arabidopsis. However, in F78Ri plants, the expression of JA-responsive pathogenesis-related genes, PBZ1 and PR1b, was induced after inoculation with M. grisea, although the JA-mediated wound response was suppressed. Furthermore, the application of JA methyl ester had no significant effect on the enhanced resistance in F78Ri plants. Taken together, our results indicate that, although suppression of fatty acid desaturases involves the concerted action of varied oxylipins via diverse metabolic pathways, 18:3 or 18:3-derived oxylipins, except for JA, may contribute to signaling on defense responses of rice to M. grisea infection.. |
| 29. | Yara, A., Yaeno, T., Hasegawa, T., Seto, H., Montillet, J.-L., Kusumi, K., Seo, S. and Iba, K. , Disease resistance against Magnaporthe grisea is enhanced in transgenic rice with suppression of ω-3 fatty acid desaturases., Plant and Cell Physiology, 48: 1263-1274, 2007.08. |
| 30. | Kusumi, K., Yaeno, T., Kojo, K., Hirayama, M., Hirokawa, D., Yara, A., and Iba, K., The role of salicylic acid in the glutathione-mediated protection against photo-oxidative stress in rice, Physiol. Plant., 128: 651-661, 2006.12. |
| 31. | Kensuke Kusumi, Takashi Yaeno, Kaori Kojo, Mayuko Hirayama, Daishirou Hirokawa, Asanori Yara, Koh Iba, The role of salicylic acid in the glutathione-mediated protection against photooxidative stress in rice, Physiologia Plantarum, 10.1111/j.1399-3054.2006.00786.x, 128, 4, 651-661, 2006.12, Salicylic acid (SA) is known to be an essential component responsible for disease resistance in dicotyledonous plants. In rice, however, tissue contains extremely high endogenous levels of SA that do not increase after pathogen infection, suggesting that the SA has other major functions in healthy leaves. Although involvement of SA in oxidative-stress response is known in some dicotyledonous plants, antioxidative role of SA in rice is obscure. In this study, we examined the involvement of SA in the protection against oxidative stress in rice, using transgenic plants expressing the bacterial nahG gene that encodes salicylate hydroxylase, an SA-degrading enzyme. In SA-deficient NahG rice, the glutathione pool size was constitutively diminished as compared with control plants. NahG seedlings showed a delayed development phenotype, an increased susceptibility to oxidative stress and they developed light-induced lesions in their leaves without pathogen infection. Conversely, treatment with an activator of the SA-mediated defense-signaling pathway, probenazole, increased the glutathione pool size and suppressed lesion formation. These results suggest that in rice, SA has an important role in the response to high-light-induced oxidative stress, through its regulatory effects on glutathione homeostasis.. |
| 32. | Kojo, K., Yaeno, T., Kusumi, K., Matsumura, H., Fujisawa, S., Terauchi, R. and Iba, K., Regulatory mechanisms of ROI generation are affected by rice spl mutations, Plant Cell Physiol., 47:1035-1044, 2006.08. |
| 33. | Kaori Kojo, Takashi Yaeno, Kensuke Kusumi, Hideo Matsumura, Shizuko Fujisawa, Ryohei Terauchi, Koh Iba, Regulatory mechanisms of ROI generation are affected by rice spl mutations, Plant and Cell Physiology, 10.1093/pcp/pcj074, 47, 8, 1035-1044, 2006.08, Reactive oxygen intermediates (ROIs) play a pivotal role in the hypersensitive response (HR) in disease resistance. NADPH oxidase is a major source of ROI; however, the mechanisms of its regulation are unclear. Rice spl mutants spontaneously form lesions which resemble those occurring during the HR, suggesting that the mutations affect regulation of the HR. We found that spl2, spl7 and spl11 mutant cells accumulated increased amounts of H2O 2 in response to rice blast fungal elicitor. Increased accumulation of ROIs was suppressed by inhibition of NADPH oxidase in the spl cells, and was also observed in the ozone-exposed spl plants. These mutants have sufficient activities of ROI-scavenging enzymes compared with the wild type. In addition, spl7 mutant cells accumulated higher amounts of H2O2 when treated with calyculin A (CA), an inhibitor of protein phosphatase. Furthermore, spl2 mutant plants exhibited accelerated accumulation of H2O 2 and increased rates of cell death in response to wounding. These results suggest that the spl2, spl7 and spl11 mutants are defective in the regulation of NADPH oxidase, and the spl7 mutation may give rise to enhancement of the signaling pathway which protein dephosphorylation controls, while the spl2 mutation affects both the pathogen-induced and wound-induced signaling pathways.. |
| 34. | K. Kusumi, A. Yara, N. Mitsui, Y. Tozawa, and K. Iba, Characterization of a rice nuclear-encoded plastid RNA polymerase gene OsRpoTp, Plant Cell Physiol., 10.1093/pcp/pch133, 45, 9, 1194-1201, 45 (9) : 1194-1201 (2004), 2004.09. |
| 35. | H. Sugimoto, K. Kusumi, Y. Tozawa, J. Yazaki, N. Kishimoto, S. Kikuchi and K. Iba, The virescent-2 mutation inhibits translation of plastid transcripts for the plastid genetic system at an early stage of chloroplast differentiation, Plant Cell Physiol., 10.1093/pcp/pch111, 45, 8, 985-996, 45 (8) : 985-996 (2004), 2004.08. |
| 36. | A. Yara, M. Otani, K, Kusumi, O. Matsuda, T. Shimada and K. Iba, Production of transgenic Japonica rice (Oryza sativa) cultivar, Taichung 65, by the agrobacterium-mediated method, Plant Biotech., 18:305-310, 2001.12. |
| 37. | Asanori Yara, Motoyasu Otani, Kensuke Kusumi, Osamu Matsuda, Takiko Shimada, Koh Iba, Production of transgenic Japonica rice (Oryza sativa) cultivar, taichung 65, by the agrobacterium-mediated method, Plant Biotechnology, 10.5511/plantbiotechnology.18.305, 18, 4, 305-310, 2001.01, Transgenic rice plants of Taiwanese japonica rice cultivar, Taichung 65, were obtained by cocultivating scutellum calli with an Agrobacterium tumefaciens strain, EHA101, that carried a binary vector harboring the luciferase (Luc) gene driven by the CaMV35S promoter. The transformation efficiency of Taichung 65 was similar to that obtained by the methods routinely used for a transformable cultivar, Notohikari. There was no correlation between the length of the culture period (21 to 30 days) and the transient transformation efficiency in Taichung 65 and in Notohikari. In the T0- and T1-generations, the transgenes were integrated and stably expressed, indicating that the transgene was inherited to the next generation. The copy number of integrated transgene varied from one to three in the T1-transformants, which was confirmed by Southern blot analysis. Moreover, approximately 60% of the T1-transformants of Taichung 65 showed the LUC-positive phenotype. These results suggest that, in addition to Notohikari, Taichung 65 is a practical, transformable cultivar.. |
| 38. | K. Kusumi, H. Komori, H. Satoh and K. Iba, Characterization of a zebra mutant of rice with increased susceptibility to light stress, Plant Cell Physiol., 41, 2, 158-164, 41:158-164, 2000.02. |
| 39. | N. Araki, K. Kusumi, K. Masamoto, Y. Niwa and K. Iba, Temperature-sensitive Arabidopsis mutant defective in 1-deoxy-D-xylulose 5-phosphate synthase within the plastid non-mevalonate pathway of isoprenoid biosynthesis, Physiol. Plant., 10.1034/j.1399-3054.2000.108001019.x, 108, 1, 19-24, 108:19-24, 2000.01. |
| 40. | Kensuke Kusumi, Hisayo Komori, Hikaru Satoh, Koh Iba, Characterization of a zebra mutant of rice with increased susceptibility to light stress, Plant and Cell Physiology, 10.1093/pcp/41.2.158, 41, 2, 158-164, 2000.01, The rice zebra mutant TCM248 is a single recessive mutant. This mutant develops transverse-striped leaves with green and white sectors under alternate light/dark growth conditions. Mutants that were grown under a higher light intensity during the light period showed a more intense striped phenotype. The white tissues contained abnormal chloroplasts with few internal membrane structures, while the green tissues in the mutants contained normal chloroplasts. The white tissue contained only trace amounts of Chls and carotenoids, and mRNA accumulation of nuclear genes encoding chloroplast proteins (rbcS, cab) was strongly suppressed compared to that in the wild type plants. A series of growth condition shift experiments demonstrated that the mutant displayed the striped phenotype only if it was exposed to the alternate light/dark growth conditions during a limited stage of early leaf development. These data suggest that the zebra gene is involved in the acquisition of photoprotective capacity of the plants and that this gene functions at an early stage of chloroplast differentiation.. |
| 41. | Horiguchi, G., Kawakami, N., Kusumi, K., Kodama, H. and Iba, K. , Developmental ragulation of genes for microsome and plastid omega-3 fatty acid desaturases in wheat (Triticum aestivum L.). , Plant Cell Physiol. , 39, 5, 540-544, 1998.11. |
| 42. | G. Horiguchi, N. Kawakami, K. Kusumi, H. Kodama and K. Iba, Developmental ragulation of genes for microsome and plastid ω-3 fatty acid desaturases in wheat (Triticum aestivum L.), Plant Cell Physiol., 39, 5, 540-544, 39:540-544, 1998.05. |
| 43. | Gorou Horiguchi, Naoto Kawakami, Kensuke Kusumi, Hiroaki Kodama, Koh Iba, Developmental regulation of genes for microsome and plastid ω-3 fatty acid desaturases in wheat (Triticum aestivum L.), Plant and Cell Physiology, 10.1093/oxfordjournals.pcp.a029402, 39, 5, 540-544, 1998.05, ω-3 fatty acid desaturase is responsible for the production of α-linolenic acid, which is a major fatty acid constituent of membrane lipids in higher plants. In this study, the developmental regulation of the expression of two genes, TaFAD3 and TaFAD7, which respectively encode the microsome and plastid ω-3 fatty acid desaturases in wheat, was analyzed by in situ hybridization. The transcript of TaFAD3 was abundant in the apical meristems of both shoot and root, the leaf primordium, the leaf basal intercalary meristem, and the root vascular tissues. The level of TaFAD3 mRNA was also high in embryos developing after flowering. In mature embryos, however, it was low, whereas in germinating embryos it was once again high. These results suggest that the TaFAD3 mRNA is accumulated at a high level in cells that have an ability to proliferate. In contrast, the TaFAD7 mRNA level was very low in the shoot apical meristem, leaf primordium and leaf basal intercalary meristem, but a large accumulation of TaFAD7 mRNA was found in green tissues, suggesting that the accumulation of this mRNA is associated with chloroplast development. Although the spatial distribution patterns of TaFAD3 and TaFAD7 mRNAs clearly differed, the enhanced accumulation of the TaFAD3 and TaFAD7 mRNAs was linked with the active membrane biogenesis required for cell division and with the development of the thylakoid system in plastids, respectively.. |
| 44. | Kensuke Kusumi, H. Inada, Koh Iba, New transcriptional apparatus in plastids of higher plants, Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme, 43, 3, 216-225, 1998.01. |
| 45. | Hitoshi Inada, Kensuke Kusumi, Mitsuo Nishimura, Koh Iba, Specific expression of the chloroplast gene for RNA polymerase (rpoB) at an early stage of leaf development in rice, Plant and Cell Physiology, 10.1093/oxfordjournals.pcp.a028936, 37, 2, 229-232, 1996.03, The rpoB gene for the β subunit of rice chloroplast RNA polymerase was found to be highly expressed in unexpanded immature levels that contained proplastids, indicating the specific expression of rpoB at early stage of chloroplast development. A putative transcription start site (tss) was identifield, but the 5′ upstream region of the tss had no sequences resembling typical -35 and -10 elements. A palindromic sequence and high AT-content were recognized.. |
| 46. | Kensuke Kusumi, Hitoshi Inada, Shun Ichiro Kawabata, Koh Iba, Mitsuo Nishimura, Chlorophyll deficiency caused by a specific blockage of the C5-pathway in seedlings of virescent mutant rice, Plant and Cell Physiology, 35, 3, 445-449, 1994.12, Temperature-sensitive mutants of rice, designated virescent (v1, v2 and v3), develop chlorophyll-deficient leaves at a restrictive temperature (20°C) but develop nearly normal green leaves at a permissive temperature (30°C). Analysis of the chlorophyll biosynthetic pathway in the virescent mutants indicated that the chlorophyll deficiency at the restrictive temperature was due to specific blockage of the C5-pathway. Northern analysis suggested that the chlorophyll deficiency in the virescent mutants was caused by specific inhibition of the expression of chloroplast tRNAGlu.. |
| 47. | Kensuke Kusumi, Hiroyuki Arata, Ikuko Iwasaki, Mitsuo Nishimura, Regulation of PEP-carboxylase by biological clock in a CAM plant, Plant and Cell Physiology, 35, 2, 233-242, 1994.12, The endogenous circadian rhythm in a crassulacean acid metabolism (CAM) plant Graptopetalum paraguayense was investigated. Phosphoenolpyruvate carboxylase (PEP-C) takes two forms: the malate-sensitive day form and the malate-insensitive night form. We monitored the state of PEP-C by measuring the sensitivity to malate as a parameter of the circadian rhythm. We also measured vacuolar pH and malate concentration, and contents of oxaloacetate, pyruvate and phosphoenolpyruvate (PEP).A free-running circadian oscillation was observed under continuous dim light (5 klux) after 12 h/12 h light/dark cycles at 20°C. The period of the rhythm was about 20 h. Under continuous light (18 klux), the rhythm was less clear but the length of the period was not affected. On the other hand, the rhythms of the vacuolar pH and the malate concentration were evident under the continuous light, but were not clear under the continuous dim light. The rhythm disappeared in continuous darkness. The content of PEP changed simultaneously with the transformation of PEP-C during the normal day-night cycles and under the continuous light, but stayed at a low level under the continuous dim light. This indicated that the transformation of PEP-C was not sufficient to maintain the rhythm in the carbon metabolism.Shift of the timing of the start or end of the dark period prior to the continuous illumination shifted the phase of the PEP-C rhythm without changing the period length significantly. At 30°C, the rhythm of PEP-C was less clear, but the period length was not affected.These results suggest that the biological clock controls CO2 uptake and day-night CAM cycle through regulation of PEP-C transformation.. |
| 48. | Kusumi, K., Arata, H., Iwasaki, I. and Nishimura, M., Regulation of PEP-carboxylase by biological clock in a CAM plant. , Plant Cell Physiol. , 35, 2, 233-242, 1994.02. |
| 49. | Hiroyuki Arata, Ikuko Iwasaki, Kensuke Kusumi, Mitsuo Nishimura, Thermodynamics of malate transport across the tonoplast of leaf cells of CAM plants, Plant and Cell Physiology, 33, 7, 873-880, 1992.10, The electrochemical potential difference for each dissociation state of malic acid across the tonoplast of leaf cells was examined in two CAM plants, Graptopetalum paraguayense and Kalanchoë daigremontiana. The concentration of malic acid in each dissociation state was estimated from an analysis of pH and concentrations of ionic species that included calcium, malate and isocitrate. The vacuoles contained 30-40 mM isocitrate and 50-70 mM calcium in G. paraguayense, and 20-30 mM isocitrate and 70-100 mM calcium in K. daigremontiana. For the calculation of the pattern of dissociation of malic acid, the formation of chelates of calcium with malate and isocitrate, which have different stability constants depending on the dissociation of the acids, were also taken into consideration. The vacuolar concentrations of the divalently dissociated form of malic acid (mal2- were 4-7 mM and 1-3 mM in G. paraguayense and in K. daigremontiana, respectively. To obtain information about the cytoplasmic concentration of malate, the apparent inhibition constant for malate of phosphoenolpyruvate carboxylase was measured. It was about 330 μM in the dark period and 60 μM in the light period. Considering an inside-positive membrane potential, we conclude that mal2- can be taken up passively into the vacuole during the dark period and can be released passively from the vacuole during the light period. Two types of channel (the "SV-type" channel and a novel "MU-type" channel) which we found recently in G. paraguayense [Iwasaki et al. (1992) Plant Physiol. 98: 1494] are probably involved in the uptake and the release of malate in the diurnal CAM rhythm. The existence of a large pH-buffering capacity due to isocitric acid in the vacuole allows the accumulation of a large amount of malic acid during the diurnal CAM rhythm.. |
| 50. | Kusumi, K., Fuse, T., Nishiuchi, T., Iba, K. and Nishimura, M. , Temperature-sensitive expression of chloroplast tRNA(Glu) in virescent mutants of rice. , Research in Photosynthesis , 3, 1, 182-182, 1992.02. |
| 51. | Arata, H., Iwasaki, I., Kusumi, K. and Nishimura, M. , Thermodynamics of malate transport across the tonoplast of leaf cells of CAM plants. , Plant Cell Physiol. , 33, 7, 873-880, 1992.02. |
| 52. | Kusumi, K., Arata, H., Iwasaki, I. and Nishimura, M., Circadian rhythms in the regulation of phosphoenolpyruvate carboxylase from a CAM plant. , Research in Photosynthesis , 3, 835-838, 1992.02. |
| 53. | Iwasaki, I., Arata, H., Kusumi, K. and Nishimura, M , Regulation of the vacuolar ion channels and cellular compartments of leaf cells of CAM plants. , Research in Photosynthesis , 3, 899-902, 1992.02. |
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