Hirokazu Tsukaya

The University of Tokyo, Tōkyō, Japan

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Publications (154)600.39 Total impact

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    ABSTRACT: Plant shoot organs such as stems, leaves, and flowers are derived from specialized groups of stem cells organized at the shoot apical meristem (SAM). Organogenesis involves two major processes, namely, cell proliferation and differentiation, whereby the former contributes to increasing the cell number and the latter involves substantial increases in cell volume through cell expansion. Coordination between the above processes in time and space is essential for proper organogenesis. To identify regulatory factors involved in proper organogenesis, heavy-ion beam-irradiated de-etiolated (det) 3-1 seeds have been used to identify striking phenotypes in the A#26-2; det3-1 mutant. In addition to the stunted plant stature mimicking det3-1, the A#26-2; det3-1 mutant exhibited stem thickening, increased floral organ number, and a fruit shape reminiscent of clavata (clv) mutants. DNA sequencing analysis demonstrated that A#26-2; det3-1 harbors a mutation in the CLV3 gene. Importantly, A#26-2; det3-1 displayed cracks that randomly occurred on the main stem with a frequency of ~50%. Furthermore, the double mutants clv3-8 det3-1, clv1-4 det3-1, and clv2-1 det3-1 consistently showed stem cracks with frequencies of ~97%, 38%, and 35%, respectively. Cross sections of stems further revealed an increase in vascular bundle number, cell number, and size in the pith of clv3-8 det3-1 compared to det3-1. These findings suggest that the stem inner volume increase due to clv mutations exerts an outward mechanical stress; that in a det3-1 background (defective in cell expansion) resulted in cracking of the outermost layer of epidermal cells.
    Plant and Cell Physiology 09/2014; In press. · 4.98 Impact Factor
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    ABSTRACT: In compound leaves, leaflet primordia are initiated directionally along the lateral sides. Our understanding of the molecular basis of leaflet initiation has improved, but the regulatory mechanisms underlying spatio-temporal patterns remain unclear. In this study, we investigated the mechanisms of acropetal (from the base to the tip) progression of leaflet initiation in Eschscholzia californica. We established an ultraviolet-laser ablation system to manipulate compound-leaf development. Local ablation at the leaflet incipient site generated leaves with asymmetric morphology. In the majority of cases, leaflets that were initiated on the ablated sides shifted apically. Finite time-course observation revealed that the timing of leaflet initiation was delayed, but the distance from the leaf tip did not decrease. These results were suggestive of the local spacing mechanism in leaflet initiation, whereby the distance from the leaf tip and adjacent pre-existing leaflet determines the position of leaflet initiation. To understand how such a local patterning mechanism generates a global pattern of successive leaflet initiation, we assessed the growth rate gradient along the apical-basal axis. Our time-course analysis revealed differential growth rates along the apical-basal axis of the leaf, which can explain the acropetal progression of leaflet initiation. We propose that a leaflet is initiated at a site where the distances from pre-existing leaflets and the leaf tip are sufficient. Furthermore, the differential growth rate may be a developmental factor underlying the directionality of leaflet initiation.
    Planta 04/2014; · 3.38 Impact Factor
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    Hirokazu Tsukaya
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    ABSTRACT: Recent accumulation of our knowledge on basic leaf development mechanisms in model angiosperm species has allowed us to pursue evolutionary development (evo/devo) studies of various kinds of leaf development. As a result, unexpected findings and clues have been unearthed aiding our understanding of the mechanisms involved in the diversity of leaf morphology, although the covered remain limited. In this review, we highlight recent findings of diversified leaf development in angiosperms.
    Current opinion in plant biology 02/2014; 17C:103-109. · 10.33 Impact Factor
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    ABSTRACT: Plant development has been evaluated at various developmental stages, from the early steps of embryogenesis to flowering. In most reports, transcription factors have been thought to play a master regulatory role in the complex networks orchestrating organogenesis. Although these efforts have increased our understanding of several major developmental pathways, our understanding of the relationships between metabolism and development remains limited. Recently, we identified a straightforward relationship linking carbohydrate metabolism and organogenesis. We found that plant development, particularly the reactivation of cell cycling after germination and the transition from heterotrophic to autotrophic growth, are highly dependent on sucrose availability. In the case of Arabidopsis thaliana, an oilseed species, we characterized the importance of cytosolic inorganic pyrophosphate hydrolysis for the success of the above transition and appropriate execution of postembryonic developmental programs. While this unprecedented and unique discovery has addressed fundamental issues concerning the biological role of the proton-pyrophosphatase (H+-PPase), it has also raised questions regarding the link between metabolism and development. Here, we summarize our present knowledge of key steps in the mobilization of storage lipids and their impact together with H+-PPase during the heterotrophic-autotrophic growth transition.
    Plant Morphology. 01/2014; 26:45-51.
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    ABSTRACT: The Cayratia japonica-Cayratia tenuifolia species complex (Vitaceae) is distributed from temperate to tropical East Asia, Southeast Asia, India, and Australia. The spatiotemporal diversification history of this complex was assessed through phylogenetic and biogeographic analyses. Maximum parsimony, neighbor-joining, and maximum likelihood methods were used to analyze sequences of one nuclear (AS1) and two plastid regions (trnL-F and trnC-petN). Bayesian dating analysis was conducted to estimate the divergence times of clades. The likelihood method LAGRANGE was used to infer ancestral areas. The Asian C. japonica and C. tenuifolia should be treated as an unresolved complex, and Australian C. japonica is distinct from the Asian C. japonica-C. tenuifolia species complex and should be treated as separate taxa. The Asian C. japonica-C. tenuifolia species complex was estimated to have diverged from its closest relatives during the Late Eocene (35.1 million years ago [Ma], 95% highest posterior densities [HPD] = 23.3-47.3 Ma) and most likely first diverged in mid-continental Asia. This complex was first divided into a northern clade and a southern clade during the middle Oligocene (27.3 Ma; 95% HPD = 17.4-38.1 Ma), which is consistent with a large southeastward extrusion of the Indochina region relative to South China along the Red River. Each of the northern and southern clades then further diverged into multiple subclades through a series of dispersal and divergence events following significant geological and climatic changes in East and Southeast Asia during the Miocene. Multiple inter-lineage hybridizations among four lineages were inferred to have occurred following this diversification process, which caused some Asian lineages to be morphologically cryptic.
    Molecular Phylogenetics and Evolution 01/2014; · 4.07 Impact Factor
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    ABSTRACT: To gain more insight into the physiological function of nitrogen dioxide (NO2 ), we investigated the effects of exogenous NO2 on growth in Arabidopsis thaliana. Plants were grown in air without NO2 for 1 wk after sowing and then grown for 1-4 wk in air with (designated treated plants) or without (control plants) NO2 . Plants were irrigated semiweekly with a nutrient solution containing 19.7 mM nitrate and 10.3 mM ammonium. Five-week-old plants treated with 50 ppb NO2 showed a ≤ 2.8-fold increase in biomass relative to controls. Treated plants also showed early flowering. The magnitude of the effects of NO2 on leaf expansion, cell proliferation and enlargement was greater in developing than in maturing leaves. Leaf areas were 1.3-8.4 times larger on treated plants than corresponding leaves on control plants. The NO2 -induced increase in leaf size was largely attributable to cell proliferation in developing leaves, but was attributable to both cell proliferation and enlargement in maturing leaves. The expression of different sets of genes for cell proliferation and/or enlargement was induced by NO2 , but depended on the leaf developmental stage. Collectively, these results indicated that NO2 regulates organ growth by controlling cell proliferation and enlargement.
    New Phytologist 12/2013; · 6.74 Impact Factor
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    ABSTRACT: Compensation refers to an increase in cell size when the cell number is significantly decreased due to the mutation or gain of function of a gene that negatively affects the cell cycle. Given the importance of coordinated growth during organogenesis in both animal and plant systems, compensation is important to understand the mechanism of size regulation. In leaves, cell division precedes cell differentiation (which involves cell expansion); therefore, a decrease in cell number triggers enhanced cell expansion (compensated cell enlargement; hereafter, CCE). Functional analyses of genes for which a loss or gain of function triggers compensation have increased our understanding of the molecular mechanisms underlying the decrease in cell number. Nevertheless, the mechanisms that induce enhanced cell expansion (the link between cell cycling and expansion), as well as the cellular machinery mediating CCE, have not been characterized. We recently characterized an important pathway involved in cell enlargement in KRP2-overexpressing plants. Here, we discuss the potential axial role of plant KRPs in triggering enlargement in cells with meristematic features.
    Plant signaling & behavior 12/2013; 8(11).
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    ABSTRACT: Compensation refers to an increase in cell size when the cell number is significantly decreased due to the mutation or gain of function of a gene that negatively affects the cell cycle. Given the importance of coordinated growth during organogenesis in both animal and plant systems, compensation is important to understand the mechanism of size regulation. In leaves, cell division precedes cell differentiation (which involves cell expansion); therefore, a decrease in cell number triggers enhanced cell expansion (compensated cell enlargement; hereafter, CCE). Functional analyses of genes for which a loss or gain of function triggers compensation have increased our understanding of the molecular mechanisms underlying the decrease in cell number. Nevertheless, the mechanisms that induce enhanced cell expansion (the link between cell cycling and expansion), as well as the cellular machinery mediating CCE, have not been characterized. We recently characterized an important pathway involved in cell enlargement in KRP2-overexpressing plants. Here, we discuss the potential axial role of plant KRPs in triggering enlargement in cells with meristematic features.
    Plant signaling & behavior 11/2013; In press.
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    ABSTRACT: Leaves are determinate organs; hence, precise control of cell proliferation and post-mitotic cell expansion is essential for their growth. A defect in cell proliferation often triggers enhanced post-mitotic cell expansion in leaves. This phenomenon is referred to as 'compensation'. Several lines of evidence from studies on compensation have shown that cell proliferation and post-mitotic cell expansion are coordinately regulated during leaf development. Therefore, compensation has attracted much attention to the mechanisms for leaf growth. However, our understanding of compensation at the subcellular level remains limited because studies of compensation have focused mainly on cellular-level phenotypes. Proper leaf growth requires quantitative control of subcellular components in association with cellular-level changes. To gain insight into the subcellular aspect of compensation, we investigated the well-known relationship between cell area and chloroplast number per cell in compensation-exhibiting lines, and asked whether chloroplast proliferation is modulated in response to the induction of compensation. We first established a convenient and reliable method for observation of chloroplasts in situ. Using this method, we analyzed Arabidopsis thaliana mutants fugu5 and angustifolia3 (an3), and a transgenic line KIP-RELATED PROTEIN2 overexpressor (KRP2 OE), which are known to exhibit typical features of compensation. We here showed that chloroplast number per cell increased in the subepidermal palisade tissue of these lines. We analyzed tetraploidized wild type, fugu5, an3 and KRP2 OE, and found that cell area itself, but not nuclear ploidy, is a key parameter that determines the activity of chloroplast proliferation. In particular, in the case of an3, we uncovered that promotion of chloroplast proliferation depends on the enhanced post-mitotic cell expansion. The expression levels of chloroplast proliferation-related genes are similar to or lower than that in the wild type during this process. This study demonstrates that chloroplast proliferation is promoted in compensation-exhibiting lines. This promotion of chloroplast proliferation takes place in response to cell-area increase in post-mitotic phase in an3. The expression of chloroplast proliferation-related genes were not promoted in compensation-exhibiting lines including an3, arguing that an as-yet-unknown mechanism is responsible for modulation of chloroplast proliferation in these lines.
    BMC Plant Biology 09/2013; 13(1):143. · 4.35 Impact Factor
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    ABSTRACT: Decreased cell numbers during leaf development often triggers increased cell size, a phenomenon called compensation. In compensation-exhibiting mutants, the unusually high cell expansion activity occurs through two different mechanisms during the post-mitotic stage of leaf development, except in the KIP-RELATED PROTEIN 2 over-expressing line (KRP2 o/e), whose cell sizes are twofold greater during proliferative growth. However, the molecular basis of compensated cell expansion (CCE) has not been characterized. The det3-1 mutant has a mutation in the C-subunit of the V-ATPase complex that causes a 50% decrease in its activity and cell size. To determine the contribution of V-ATPase activity to CCE, the cellular phenotypes of double mutants between det3-1 and compensation-exhibiting fugu5-1, an3-4, fas1-5 and KRP2 o/e were analyzed in detail. Interestingly, while decreased V-ATPase activity caused by det3-1 did not suppress CCE in fugu5-1, fas1-5 and an3-4, CCE in KRP2 o/e was suppressed totally. Furthermore, measurements revealed that the activity and quantity of the A subunit of the V-ATPase complex were significantly increased in the shoots of KRP2 o/e plants. Importantly, the unusually increased size of actively dividing KRP2 o/e cells was restored to normal in the det3-1 background. Taken together, our data strongly suggest that CCE in KRP2 o/e, but not in other compensation-exhibiting mutants, occurs exclusively through the increase of V-ATPase activity.
    Plant and Cell Physiology 09/2013; · 4.98 Impact Factor
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    ABSTRACT: • Premise of the study: On a compound leaf, leaflet primordia are repetitively formed along the apical-basal axis, with the direction varying among taxa. Why and how the directions vary among species is yet to be solved, although a change in a single factor was proposed to cause the variation. In this study, we compared two species in the Papaveraceae with different directions of leaflet initiation, Chelidonium majus subsp. asiaticum (basipetal) and Eschscholzia californica (acropetal). Because E. californica has been studied in some detail, we focused on C. majus and asked how basipetal pattern is achieved.• Methods: Since only immature leaf primordial tissue has leaflet-generating competency, we performed histological and gene expression analyses on markers of the tissue maturation state. In addition, we performed a time-course analysis of leaf primordial growth.• Key results: Quantitative reverse transcription-PCR analysis demonstrated that a putative regulator of tissue maturation in C. majus, the CINCINNATA homolog, had higher expression in apical parts than in basal parts during the organogenetic phase. In contrast, expression of the CIN homolog was not elevated in either the apical or basal parts in E. californica during the organogenetic phase.• Conclusions: In C. majus, apical parts of leaf primordia have already lost leaflet-generating competency during the organogenetic phase. We propose that precocious progression of the maturation process instructs basipetal progression of leaflet initiation in C. majus. This is not the mirror image of data on E. californica, which shows the opposite direction in leaflet formation, indicating that variation in direction is not attributable to a change in a single factor.
    American Journal of Botany 05/2013; · 2.59 Impact Factor
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    ABSTRACT: During leaf development, a decrease in cell number often triggers an increase in cell size. This phenomenon, called compensation, suggests that some system coordinates cell proliferation and cell expansion but how this is mediated at the molecular level is still unclear. The fugu2 mutants in Arabidopsis thaliana exhibit typical compensation phenotypes. Here we report that the FUGU2 gene encodes FASCIATA1 (FAS1), the p150 subunit of chromatin assembly factor-1 (CAF-1). To uncover how fas1 mutation induces compensation, we performed microarray analyses and found that many genes involved in the DNA damage response are up-regulated in fas1. Our genetic analysis further showed that activation of the DNA damage response and accompanying decrease of cell number in fas1 depend on ATAXIA TELANGIECTASIA MUTATED (ATM) but not on ATM AND RAD3 RELATED (ATR). Kinematic analysis suggested that the delay in the cell cycle leads to a decrease in cell number in fas1 and that loss of ATM partially restores this phenotype. Consistently, both cell size phenotypes and high ploidy phenotypes of fas1 are also suppressed by atm, supporting that ATM-dependent DNA damage response leads to these phenotypes. Altogether, these data suggest that ATM-dependent DNA damage response acts as an upstream trigger in fas1 to delay the cell cycle and promote an entry into the endocycle, resulting in compensated cell expansion.
    Plant physiology 04/2013; · 6.56 Impact Factor
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    ABSTRACT: Coordinated proliferation between clonally distinct cells via inter-cell-layer signaling largely determines the size and shape of plant organs [1-4]. Nonetheless, the signaling mechanism underlying this coordination in leaves remains elusive because of a lack of understanding of the signaling molecule (or molecules) involved. ANGUSTIFOLIA3 (AN3, also called GRF-INTERACTING FACTOR1) encodes a putative transcriptional coactivator with homology to human synovial sarcoma translocation protein [5-7]. AN3 transcripts accumulate in mesophyll cells but are not detectable in leaf epidermal cells [8]. However, we found here that in addition to mesophyll cells [5, 6], epidermal cells of an3 leaves show defective proliferation. This spatial difference between the accumulation pattern of AN3 transcripts and an3 leaf phenotype is explained by AN3 protein movement across cell layers. AN3 moves into epidermal cells after being synthesized within mesophyll cells and helps control epidermal cell proliferation. Interference with AN3 movement results in abnormal leaf size and shape, indicating that AN3 signaling is indispensable for normal leaf development. AN3 movement does not require type II chaperonin activity, which is needed for movement of some mobile proteins [9]. Taking these findings together, we present a novel model emphasizing the role of mesophyll cells as a signaling source coordinating proliferation between clonally independent leaf cells.
    Current biology: CB 04/2013; · 10.99 Impact Factor
  • Hirokazu Tsukaya, Hiroshi Okada
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    ABSTRACT: Irrespective of the fact that Borneo is one of the richest areas for mycoheterotrophic plants, only two species of the genus Lecanorchis Blume, a mycoheterotrophic genus of Orchidaceae, have been reported from Borneo. On three occasions during recent botanical surveys in Betung Kerihun National Park, we found an undescribed species of Lecanorchis. Here, we provide a detailed morphological account of this new species, Lecanorchis betung-kerihunensis. We also provide a key to the species of Lecanorchis in Borneo.
    Systematic Botany 03/2013; 38(1):69-74. · 1.29 Impact Factor
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    ABSTRACT: Shade avoidance response (S.A.R) is regulated by light and circadian clock. Circadian clock controls S.A.R by the transcriptional regulation of positive regulators of S.A.R, PIF4 and PIF5, to prevent plants from responding to 'light' of dark period. Thus, in many cases, deficits in circadian clock appear in abnormalities of hypocotyl and/or petiole elongation. Previously, interesting phenomena were reported that the triple mutants of PSEUDO RESPONSE REGULATORS 9, 7 and 5, which are clock components, show longer petioles and smaller leaves under light/dark cycle than those under continuous lighting. These S.A.R-like phenotypes cannot be explained by their hyposensitivity to red light. We demonstrated detailed analyses of this mutant to reveal the leaf-specific S.A.R regulated by circadian clock. Expression analyses of S.A.R-related genes suggested that PRR5 functions as a repressor of S.A.R. Morphological analyses of leaves under different light condition revealed that PRR5 is involved in the inhibition of leaf expansion in S.A.R.
    Plant signaling & behavior 01/2013; 8(4).
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    Hokuto Nakayama, Takahiro Yamaguchi, Hirokazu Tsukaya
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    ABSTRACT: It has been suggested that modification and co-option of existing gene regulatory networks (GRNs) play an important role in the morphological diversity. In plants, leaf development is one of active research areas, and the basic GRN for leaf development is beginning to be understood. Moreover, leaves show wide variation in their form, and some of this variation is thought to be the result of adaptation. Thus, leaves and leaf-like organs are an emerging and interesting model to reveal how existing GRNs give rise to novel forms and architectures during evolution. In this review, we highlight recent findings in evo-devo studies, especially on Juncus unifacial leaves, which are composed of lamina with abaxialized identities, and Asparagus cladodes, which are leaf-like organs at the axils of scale leaves. Based on these studies, we discuss how flat structures have evolved and morphologically diversified in shoot systems of monocot species, focusing on the modification and co-option of GRN for leaf development.
    Frontiers in Plant Science 01/2013; 4:248. · 3.60 Impact Factor
  • Hirokazu Tsukaya
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    ABSTRACT: Leaves are the most important organs for plants. Without leaves, plants cannot capture light energy or synthesize organic compounds via photosynthesis. Without leaves, plants would be unable perceive diverse environmental conditions, particularly those relating to light quality/quantity. Without leaves, plants would not be able to flower because all floral organs are modified leaves. Arabidopsis thaliana is a good model system for analyzing mechanisms of eudicotyledonous, simple-leaf development. The first section of this review provides a brief history of studies on development in Arabidopsis leaves. This history largely coincides with a general history of advancement in understanding of the genetic mechanisms operating during simple-leaf development in angiosperms. In the second section, I outline events in Arabidopsis leaf development, with emphasis on genetic controls. Current knowledge of six important components in these developmental events is summarized in detail, followed by concluding remarks and perspectives.
    The Arabidopsis Book 01/2013; 11:e0163.
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    Hirokazu Tsukaya
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    ABSTRACT: Ploidy level affects cell size in many organisms, and ploidy-dependent cell enlargement has been used to breed many useful organisms. However, how polyploidy affects cell size remains unknown. Previous studies have explored changes in transcriptome data caused by polyploidy, but have not been successful. The most naïve theory explaining ploidy-dependent cell enlargement is that increases in gene copy number increase the amount of protein, which in turn increases the cell volume. This hypothesis can be evaluated by examining whether any strains, mutants, or transgenics show the same cell size before and after a tetraploidization event. I performed this experiment by tetraploidizing various mutants and transgenics of Arabidopsis thaliana, which show a wide range in cell size, and found that the ploidy-dependent increase in cell volume is genetically regulated. This result is not in agreement with the theory described above.
    PLoS ONE 01/2013; 8(12):e83729. · 3.53 Impact Factor
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    Hirokazu Tsukaya
    EMBO Reports 12/2012; · 7.19 Impact Factor
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    Hirokazu Tsukaya
    Journal of Plant Research 12/2012; · 2.06 Impact Factor

Publication Stats

4k Citations
600.39 Total Impact Points


  • 1992–2014
    • The University of Tokyo
      • • Department of Biological Sciences
      • • Faculty of Science and Graduate School of Science
      • • Institute of Molecular and Cellular Biosciences
      • • Laboratory of Molecular Genetics
      Tōkyō, Japan
  • 2013
    • Kyoto Sangyo University
      • Faculty of Life Sciences
      Kyoto, Kyoto-fu, Japan
    • RIKEN
      Вако, Saitama, Japan
  • 2011–2013
    • Tokyo Gakugei University
      Koganei, Tōkyō, Japan
  • 2010–2012
    • Rikkyo University
      • Department of Life Science
      Tokyo, Tokyo-to, Japan
    • Hiroshima University
      • Division of Mathematical and Life Sciences
      Hiroshima-shi, Hiroshima-ken, Japan
  • 2000–2010
    • National Institute for Basic Biology
      Okazaki, Aichi, Japan
  • 2009
    • Ghent University
      • VIB Department of Plant Systems Biology
      Gent, VLG, Belgium
  • 2008
    • Yale University
      • Department of Molecular, Cellular and Developmental Biology
      New Haven, CT, United States
    • Yamagata University
      • Department of Biology
      Ямагата, Yamagata, Japan
  • 2007
    • Chubu University
      • College of Bioscience and Biotechnology
      Kasugai, Aichi-ken, Japan
  • 2006
    • National Institutes Of Natural Sciences
      Edo, Tōkyō, Japan
  • 2002–2006
    • The Graduate University for Advanced Studies
      • School of Advanced Sciences
      Miura, Kanagawa-ken, Japan
    • Nagoya University
      • Department of Biological Science
      Nagoya-shi, Aichi-ken, Japan
    • Bulgarian Academy of Sciences
      • Institute of Plant Physiology and Genetics
      Ulpia Serdica, Sofia-Capital, Bulgaria
  • 2005
    • Dong-A University
      Tsau-liang-hai, Busan, South Korea
  • 1997–2005
    • Tohoku University
      • Graduate School of Life Sciences
      Sendai-shi, Miyagi-ken, Japan