Takaya Kisugi's research while affiliated with Tohoku University and other places

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Publications (29)


Fig. 1. Proposed biosynthetic pathway of SLs. Blue, red, green, and orange letters indicate genes of rice, Arabidopsis, pea, and petunia, respectively. 2OGD, 2-oxoglutarate-dependent dioxygenase; CCD, carotenoid cleavage dioxygenase.
Fig. 2. Conversion of CLA to MeCLA by CLAMT. Enzymatic reaction was performed using CLA as a substrate in the presence of SAM. The reaction product of the cell lysate expressing the empty vector or At4g36470 (CLAMT) was analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) (Sciex X500R quadrupole time-of-flight). The MS/MS chromatogram for MeCLA (m/z 347.18 > 97.03) (Left) and MS/MS scan spectra of fragment ions between m/z 50 and 380 from a precursor ion with m/z 347.18 (Right) are shown.
Fig. 3. Analysis of the Arabidopsis clamt mutants in the No-0 background. (A) Quantitative analysis of endogenous CLA and MeCLA levels in Arabidopsis WT (No-0) and the clamt mutants. Data are the means ± SD (n = 4). (B) Shoot branching phenotype of Arabidopsis WT (No-0) and the clamt mutants. The number of axillary shoots (>5 mm) per plant of 44-d-old plants is shown as the mean ± SD (n = 9). Right panels show the pictures of the aboveground part of these plants. (C) Effect of SLs (10 μM) on axillary bud outgrowth of Arabidopsis. A solution (10 μL) containing each compound was applied to axillary buds every other day for 29 d. The number of axillary shoots (>5 mm) per 46-d-old plant is shown as the mean ± SD (n = 8). Different letters indicate significant differences at P < 0.05, Tukey's honestly significant difference (HSD).
A carlactonoic acid methyltransferase that contributes to the inhibition of shoot branching in Arabidopsis
  • Article
  • Full-text available

April 2022

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188 Reads

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44 Citations

Proceedings of the National Academy of Sciences

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Yuta Onozuka

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[...]

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Significance Strigolactones (SLs) are a group of apocarotenoid hormones, which regulates shoot branching and other diverse developmental processes in plants. The major bioactive form(s) of SLs as endogenous hormones has not yet been clarified. Here, we identify an Arabidopsis methyltransferase, CLAMT, responsible for the conversion of an inactive precursor to a biologically active SL that can interact with the SL receptor in vitro. Reverse genetic analysis showed that this enzyme plays an essential role in inhibiting shoot branching. This mutant also contributed to specifying the SL-related metabolites that could move from root to shoot in grafting experiments. Our work has identified a key enzyme necessary for the production of the bioactive form(s) of SLs.

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Fig. 1 Chemical structures of SL-related compounds and a scheme for their biosynthesis and signaling pathways. a Structures of SL-related compounds. b The scheme for the SL biosynthesis and signaling pathways. Red, blue, orange, and green characters indicate genes of Arabidopsis, rice, petunia, and pea, respectively. Black arrows indicate the biosynthetic steps, and a white arrow indicates the signaling step. (CCD; carotenoid cleavage dioxygenase)
Fig. 2 Evaluation of the AtD14-SL interaction using hydrolysis and DSF assays. a Melting temperature curves of AtD14 in the presence of various SLs and analogs. The names in red and blue denote biologically active and inactive (or weakly active) compounds, respectively. b and d, Monitoring by LC-MS/MS of the AtD14 hydrolysis reaction of GR24 (b) and CN-PMF (d). HBN; Hydroxybenzenenitrile. Data are the means ± SD (n = 3). c and e Melting temperature curves of AtD14 pre-incubated with GR24 (c) and CN-PMF (e) for indicated time period. Source data are provided as a Source Data file
Fig. 3 In vitro and in vivo functional analysis of catalytic triad mutants of AtD14. a Hydrolysis activities of catalytic triad mutants of AtD14 using 1 μM of 5DS as a substrate. Data are the means ± SD (n = 3). The control reaction (Cont.) is for MBP protein only. b No. of axillary shoots (over 5 mm) of Arabidopsis transgenic plants expressing each catalytic triad mutant of AtD14 in the atd14-2 mutant background. Data are the means ± SD (n = 5-10, Different letters indicate significant differences at P < 0.05 with Tukey-kramer multiple comparison test.). c Phenotypes of Arabidopsis transgenic plants expressing each catalytic triad mutant of AtD14. Mature 50 days old plants phenotypes (upper panel) and leaf morphology phenotypes of 25 days old plants (lower panel) are shown. Scale bars = 5 cm (upper panel), 1 cm (lower panel). d Y2H analysis of the interaction between SMXL7 and each catalytic triad mutant of AtD14. Yeast transformants were spotted onto the control medium (SD−Leu/−Trp (−TL)) and selective medium (SD−Leu/−Trp/−His (−TLH)) in the absence or presence of SLs (10 μM rac-MeCLA or 10 μM 5DS). Control (Cont.) is acetone only. e Shoot branching inhibition assays of the Arabidopsis transgenic lines expressing AtD14 WT and AtD14 D218A , respectively, in the atd14 max4 double mutants background. The bars indicate the No. of axillary shoots (over 5 mm) in the presence (+) or absence (−) of GR24 at 5 μM (left panel) and 0.5 μM (right panel), respectively. Data are the means ± SD (n = 3-13, Different letters indicate significant differences at P < 0.05 with Tukey-kramer multiple comparison test). Source data are provided as a Source Data file
In vitro and in vivo functional analysis of OsD14R233H/AtD14R133H. a Phenotypes of 2 weeks old seedlings (left) and mature plants (right) of the rice d14-2 mutant. The white arrow in the left picture indicates the outgrowing tiller. Scale bars = 5 cm (left panel), 20 cm (right panel). b Hydrolase activities of OsD14R233H and AtD14R133H mutants using 1 μM 5DS as a substrate. Data are the means ± SD (n = 3–4). c No. of tillers of rice transgenic plants overexpressing OsD14R233H in the WT (Nipponbare) background. Empty vector expressing plants are indicated as EV. Data are the means ± SD (n = 3–5). d Phenotypes of 42 days old transgenic plants overexpressing OsD14R233H (OsD14R233HOE). Scale bars = 10 cm. e Quantitative analysis of 4DO in the root exudates (left panel) and extracts (right panel) of OsD14R233H overexpressing (OsD14R233HOE) plants. Data are the means ± SD (n = 3–4). Different letters in c and e indicate significant differences at P < 0.05 with Tukey-kramer multiple comparison test. Source data are provided as a Source Data file
A proposed working model of D14 in the SL signaling pathway. A bioactive SL molecule induces the protein conformational changes of D14, which triggers complex formation with the signaling partners. After the degradation of D53/SMXLs and transmission of the SL signal, D14 reconstructs the catalytic triad to hydrolytically decompose the bioactive SL
Strigolactone perception and deactivation by a hydrolase receptor DWARF14

January 2019

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726 Reads

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205 Citations

Nature Communications

The perception mechanism for the strigolactone (SL) class of plant hormones has been a subject of debate because their receptor, DWARF14 (D14), is an α/β-hydrolase that can cleave SLs. Here we show via time-course analyses of SL binding and hydrolysis by Arabidopsis thaliana D14, that the level of uncleaved SL strongly correlates with the induction of the active signaling state. In addition, we show that an AtD14D218A catalytic mutant that lacks enzymatic activity is still able to complement the atd14 mutant phenotype in an SL-dependent manner. We conclude that the intact SL molecules trigger the D14 active signaling state, and we also describe that D14 deactivates bioactive SLs by the hydrolytic degradation after signal transmission. Together, these results reveal that D14 is a dual-functional receptor, responsible for both the perception and deactivation of bioactive SLs.





Which are Major Players, Canonical or Non-Canonical Strigolactones?

March 2018

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131 Reads

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137 Citations

Journal of Experimental Botany

Strigolactones (SLs) can be classified into two structurally distinct groups: canonical and non-canonical SLs. Canonical SLs contain the ABCD ring system, and non-canonical SLs lack the A, B, or C ring but have the enol ether-D ring moiety which is essential for biological activities. The simplest non-canonical SL is the SL biosynthetic intermediate carlactone (CL). In plants, CL and its oxidized metabolites such as carlactonoic acid and methyl carlactonoate, are present in root and shoot tissues. In some plant species including black oat (Avena strigosa), sunflower (Helianthus annuus), and maize (Zea mays), non-canonical SLs are major germination stimulants in the root exudates. Various plant species such as tomato (Solanum lycopersicum), Arabidopsis, and poplar (Populus spp.) release carlactonoic acid into the rhizosphere. These results suggest that both canonical and non-canonical SLs are active as host recognition signals in the rhizosphere. In contrast, limited distribution of canonical SLs in the plant kingdom and structure- and stereo-specific transportation of canonical SLs from roots to shoots suggest that plant hormones inhibiting shoot branching are not canonical SLs but are rather non-canonical SLs.


Which are major players, canonical or non-canonical strigolactones?

September 2017

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25 Reads

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1 Citation

Strigolactones (SLs) can be classified into two structurally distinct groups: canonical and non-canonical SLs. Canonical SLs contain the ABCD ring system, and non-canonical SLs lack the A, B, or C ring but have the enol ether–D ring moiety which is essential for biological activities. The simplest non-canonical SL is the SL biosynthetic intermediate carlactone (CL). In plants, CL and its oxidized metabolites such as carlactonoic acid and methyl carlactonoate, are present in root and shoot tissues. In some plant species including black oat (Avena strigosa), sunflower (Helianthus annuus), and maize (Zea mays), non-canonical SLs are major germination stimulants in the root exudates. Various plant species such as tomato (Solanum lycopersicum) release carlactonoic acid, and poplar (Populus spp.) was found to exude methyl carlactonoate into the rhizosphere. These results suggest that both canonical and non-canonical SLs are active as host recognition signals in the rhizosphere. In contrast, limited distribution of canonical SLs in the plant kingdom and structure- and stereo-specific transportation of canonical SLs from roots to shoots suggest that plant hormones inhibiting shoot branching are not canonical SLs but are rather non-canonical SLs.


Table 1 . NMR spectral data for compound 1 (C 6 D 6 ) No. δ H (mult., J Hz) HMQC and DEPT δ C HMBC NOESY 
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Methyl zealactonoate, a novel germination stimulant for root parasitic weeds produced by maize

April 2017

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225 Reads

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40 Citations

Journal of Pesticide Science

One of the germination stimulants for root parasitic weeds produced by maize (Zea mays) was isolated and named methyl zealactonoate (1). Its structure was determined to be methyl (2E,3E)-4-((RS)-3,3-dimethyl-2-(3-methylbut-2-en-2-yl)-5-oxotetrahydrofuran-2-yl)-2-((((R)-4-methyl-5-oxo-2,5-dihydrofran-2-yl)oxy)methylene)but-3-enoate using by 1D and 2D NMR spectroscopy and ESI and EI–MS spectrometry. Feeding experiments with ¹³C-carlactone (CL), a biosynthetic intermediate for strigolactones, confirmed that 1 is produced from CL in maize. Methyl zealactonoate strongly elicits Striga hermonthica and Phelipanche ramosa seed germination, while Orobanche minor seeds are 100-fold less sensitive to this stimulant.


Citations (20)


... The over-branching phenotype of the SL-deficient mutant sb1 mostly disappeared when it was grafted onto a wild-type rootstock ( Figure 5A, Table S1), supporting the notion that the roots are the primary site of SL biosynthesis, which are transported acropetally to the shoots through the xylem [28,47,59]. Similar results were previously reported in other plant species [28,[59][60][61][62][63]. Over-branching, accompanied by loss of fruit yield, was also recovered in a wild-type (M82) scion grafted onto the SL-deficient rootstock of sb1 ( Figure 5 and Table S1). ...

Reference:

Tomato Mutants Reveal Root and Shoot Strigolactone Involvement in Branching and Broomrape Resistance
A carlactonoic acid methyltransferase that contributes to the inhibition of shoot branching in Arabidopsis

Proceedings of the National Academy of Sciences

... Outside flowering plants, SL synthesis has been characterized in the moss Physcomitrella patens, where CCD7 and CCD8 act consecutively in CL synthesis as in angiosperms (Proust et al, 2011;Decker et al, 2017). There is some uncertainty about which strigolactones are ultimately synthesised by P. patens, with recent analysis suggesting only CL is produced, consistent with the lack of MAX1 orthologue in this species (Decker et al, 2017;Yoneyama et al, 2017). In general, conclusions regarding SL synthesis outside the angiosperms are based on very limited sampling of sequences. ...

Which are major players, canonical or non-canonical strigolactones?
  • Citing Preprint
  • September 2017

... SLs are a group of terpenoid phytohormones responsible for a wide range of developmental processes, especially the branching of shoots, and responses to stresses (Korek and Marzec 2023). SLs are sensed by their receptor DWARF14 (AtD14/OsD14) in Arabidopsis and rice, respectively (Seto et al. 2019). The activated receptor, in turn, interacts with an F-box protein MORE AXILLARY GROWTH 2 (MAX2) in Arabidopsis, an ortholog of DWARF3 (D3) in rice, from the SCF complex SCF MAX2 , and SL repressors SUPPRESSORS OF MAX2 1-LIKE 6/7/8 (SMXL6/7/8) in Arabidopsis or D53 in rice, reducing the stability of SMXL6/7/8 (D53) proteins (Jiang et al. 2013, Zhou et al. 2013a). ...

Reference:

pcae047-1
Strigolactone perception and deactivation by a hydrolase receptor DWARF14

Nature Communications

... Another intriguing question in the biology of SLs is related to their structural diversity ( Table 1 provides a summary of identified SLs in different species) [12,19,24,29,31,36,[41][42][43][44][45][46]51,52,65,66,73,74,[79][80][81][82][83][84][85][86][87][88][89][90][91][92] and the structure-biological function relationship: do all SLs fulfill the same hormonal functions in planta and act as rhizospheric signals [93][94][95][96]? Both SL classes are assumed to height, stem thickness, and leaf senescence, whereas they inhibit shoot branching and the growth of adventitious and lateral roots [10,56] in the non-arbuscular mycorrhiza (AM) host arabidopsis. ...

Which are Major Players, Canonical or Non-Canonical Strigolactones?
  • Citing Article
  • March 2018

Journal of Experimental Botany

... Another intriguing question in the biology of SLs is related to their structural diversity ( Table 1 provides a summary of identified SLs in different species) [12,19,24,29,31,36,[41][42][43][44][45][46]51,52,65,66,73,74,[79][80][81][82][83][84][85][86][87][88][89][90][91][92] and the structure-biological function relationship: do all SLs fulfill the same hormonal functions in planta and act as rhizospheric signals [93][94][95][96]? Both SL classes are assumed to height, stem thickness, and leaf senescence, whereas they inhibit shoot branching and the growth of adventitious and lateral roots [10,56] in the non-arbuscular mycorrhiza (AM) host arabidopsis. ...

Methyl zealactonoate, a novel germination stimulant for root parasitic weeds produced by maize

Journal of Pesticide Science

... Moreover, the catalyzing property of CL and CLA of CYP711A subfamily to canonical SLs in seed plant was exclusively detected only in rice [71]. In a previous study on faba bean, a genitor of our R_Host was used as a resistant host, and orobanchol was not detected in its root exudates [72]. Whereas, orobanchol and orobanchol acetate were detected in the root exudates of the same S_Host variety used in our study [72]. ...

Characterization of strigolactones produced by Orobanche foetida and Orobanche crenata resistant faba bean (Vicia faba L.) genotypes and effects of phosphorous, nitrogen, and potassium deficiencies on strigolactone production
  • Citing Article
  • January 2017

South African Journal of Botany

... CK transports acropetally in xylem and synthesizes locally to promote shoot branching. SL, a new phytohormone, also transports acropetally from root to shoot but it inhibits shoot branching (Gomez-Roldan et al., 2008;Xie et al., 2016). It has also recently been proved that abscisic acid (ABA) and gibberellic acid (GA) are involved in shoot branching (Yao and Scott, 2015;Charnikhova et al., 2017). ...

Structure- and stereospecific transport of strigolactones from roots to shoots

Journal of Pesticide Science

... Hence, it has been proposed that SLs are longdistance signals translocated to shoots to exert their function in inhibiting shoot tillering/ branching. Grafting experiments [61,62] and liquid chromatography-mass spectrometry (LC-MS) quantification of SLs in shoot tissues, including xylem sap [51,[63][64][65][66], demonstrate that both classes of SLs can be transported upward from the root to the shoot. However, whether a specific SL molecule is the mobile inhibitor of shoot branching in plants remains unclear. ...

Strigolactones are transported from roots to shoots, although not through the xylem

Journal of Pesticide Science

... About 50 years ago, they were known to be seed germination stimulants in root parasitic plants, e.g., Striga, Orobanche, and Phelipanche (Ruyter-Spira et al. 2013). A single plant species can produce various types of SLs, whereas in case of intraspecific varieties, blends of different kinds and amounts of SL molecules can be observed (Yoneyama et al. 2013). Though SLs are synthesized and accumulated in small quantities in roots, other parts of the plants can also produce these biomolecules. ...

Chemistry of Strigolactones: Why and How do Plants Produce so Many Strigolactones?
  • Citing Chapter
  • March 2013

... Orobanchol, a major naturally occurring SL, has been detected in the root exudate of numerous plants, including monocots like rice (Oryza sativa) (Zhang et al., 2014) and dicots such as Sonalaceae , Fabaceae (Ueno et al., 2011), Cucurbitaceae (Khetkam et al., 2014), and Linaceae (Xie et al., 2009a). Regarding SL biosynthesis, enzyme reactions catalyzed by DWARF27 (D27), carotenoid cleavage dioxygenase 7 (CCD7), and CCD8 are known to convert all-trans-b-carotene to carlactone (Alder et al., 2012). ...

7α- and 7β-Hydroxyorobanchyl acetate as germination stimulants for root parasitic weeds produced by cucumber

Journal of Pesticide Science