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Biuret toxicity induces accumulation of nitrogen-rich compounds in rice plants

Ochiai, Kumiko Nomura, Yosuke Uesugi, Asuka Matoh, Toru 京都大学 DOI:10.1007/s11104-022-05857-9

2023.04

概要

[Aims] Excess biuret, a common impurity in urea fertilizers, is toxic to plants. Little is known about the mechanisms of biuret toxicity in plants. This study aimed to investigate the accumulation of biuret and the changes in metabolites in rice (Oryza sativa) plants under biuret toxicity. [Methods] A previous study had shown that transgenic rice plants overexpressing bacterial biuret hydrolase had improved biuret tolerance. Here, we grew wild-type and bacterial biuret hydrolase-overexpressing rice plants in hydroponics at different biuret levels. Concentrations of biuret and allantoin, a nitrogenous intermediate in the purine degradation pathway, in the plants were determined. The expression levels of genes related to purine degradation and ureide metabolisms were analyzed using wild-type plants. Additionally, we performed a metabolome analysis using rice suspension cells. [Results] The biuret hydrolase-overexpressing plants did not contain biuret, whereas wild-type plants accumulated biuret in shoots in the order of mmol L−1 tissue water. The concentration of allantoin in shoots of wild-type plants under biuret toxicity was higher than those in control conditions. Inhibition of allantoinase activity by biuret was not detected, and allantoin accumulation appeared to be associated with changes in the expression of allantoinase, allantoate amidohydrolase and putative allantoin transporter genes. Furthermore, another nitrogenous compound citrulline, which is a non-protein amino acid, accumulated in rice suspension cells under biuret toxicity. [Conclusion] The accumulation of these compounds suggests that rice plants subjected to biuret toxicity need to reduce the concentration of surplus ammonium ions via synthesizing nitrogen-rich compounds.

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参考文献

455

456

Akashi K, Miyake C, Yokota A (2001) Citrulline, a novel compatible solute in drought-tolerant

457

wild watermelon leaves, is an efficient hydroxyl radical scavenger. FEBS Lett 508:438–442.

458

https://doi.org/10.1016/s0014-5793(01)03123-4

459

Aukema KG, Tassoulas LJ, Robinson SL, Konopatski JF, Bygd MD, Wackett LP (2020) Cyanuric

460

acid biodegradation via biuret: physiology, taxonomy, and geospatial distribution. Appl

461

Environ Microbiol 86:e01964-19. https://doi.org/10.1128/AEM.01964-19

462

Baba A, Hasezawa S, Syono K (1986) Cultivation of rice protoplasts and their transformation

463

mediated

464

https://doi.org/10.1093/oxfordjournals.pcp.a077122

465

Agrobacterium

spheroplasts.

Plant

Cell

Physiol

27:463–471.

Blume C, Ost J, Mühlenbruch M, Peterhänsel C, Laxa M (2019) Low CO2 induces urea cycle

466

intermediate

467

https://doi.org/10.1371/journal.pone.0210342

accumulation

Arabidopsis

thaliana.

PloS

One

14:e0210342.

468

Caldana C, Scheible WR, Mueller-Roeber B, Ruzicic S (2007) A quantitative RT-PCR platform

469

for high-throughput expression profiling of 2500 rice transcription factors. Plant Method, 3:7.

470

https://doi.org/10.1186/1746-4811-3-7

471

Cameron SM, Durchschein K, Richman JE, Sadowsky MJ, Wackett LP (2011) A new family of

472

biuret hydrolases involved in S-triazine ring metabolism. ACS Catal 2011:1075–1082.

473

https://doi.org/10.1021/cs200295n

474

Casartelli A, Melino VJ, Baumann U, Riboni M, Suchecki R, Jayasinghe NS, Mendis H,

475

Watanabe M, Erban A, Zuther E, Hoefgen R, Roessner U, Okamoto M, Heuer S (2019)

476

Opposite fates of the purine metabolite allantoin under water and nitrogen limitations in bread

477

wheat. Plant Mol Biol 99:477–497. https://doi.org/10.1007/s11103-019-00831-z

478

Collier R, Tegeder M (2012) Soybean ureide transporters play a critical role in nodule

479

development, function and nitrogen export. Plant J 72:355–367. https://doi.org/10.1111/j.1365-

480

313X.2012.05086.x

481

Desimone M, Catoni E, Ludewig U, Hilpert M, Schneider A, Kunze R, Tegeder M, Frommer W

482

B, Schumacher K (2002) A novel superfamily of transporters for allantoin and other oxo

483

derivatives of nitrogen heterocyclic compounds in Arabidopsis. Plant Cell 14:847–856.

484

https://doi.org/10.1105/tpc.010458

485

486

Duran VA, Todd CD (2012) Four allantoinase genes are expressed in nitrogen-fixing soybean.

Plant Physiol Biochem 54:149–155. https://doi.org/10.1016/j.plaphy.2012.03.002

487

Esquirol L, Peat TS, Wilding M, Lucent D, French NG, Hartley CJ, Newman J, Scott C (2018)

488

Structural and biochemical characterization of the biuret hydrolase (BiuH) from the cyanuric

489

acid catabolism pathway of Rhizobium leguminasorum bv. viciae 3841. PLoS One

490

13:e0192736. https://doi.org/10.1371/journal.pone.0192736

491

Gupta KJ, Brotman Y, Segu S, Zeier T, Zeier J, Persijn ST, Cristescu SM, Harren FJ, Bauwe H,

492

Fernie AR, Kaiser WM, Mur LA (2013) The form of nitrogen nutrition affects resistance

493

against Pseudomonas syringae pv. phaseolicola in tobacco. J Exp Bot, 64:553–568.

494

https://doi.org/10.1093/jxb/ers348

495

Hewitt EJ (1966) The composition of the nutrient solution. In: Hewitt EJ (ed) Sand and water

496

culture methods used in the study of plant nutrition. Farnham Royal Bucks, Commonwealth

497

Agricultural Bureaux, Slough, pp 190.

498

499

500

501

Huang XY, Li M, Luo R, Zhao FJ, Salt DE (2019) Epigenetic regulation of sulfur homeostasis in

plants. J Exp Bot 70:4171–4182. https://doi.org/10.1093/jxb/erz218

Irani S, Todd CD (2016) Ureide metabolism under abiotic stress in Arabidopsis thaliana. J Plant

Physiology 199:87–95. https://doi.org/10.1016/j.jplph.2016.05.011

502

503

Joshi V, Fernie AR (2017) Citrulline metabolism in plants. Amino Acids 49:1543–1559.

https://doi.org/10.1007/s00726-017-2468-4

504

Kaur H, Chowrasia S, Gaur VS, Mondal TK (2021) Allantoin: Emerging role in plant abiotic

505

stress tolerance. Plant Mol Biol Report 39:648–661. https://doi.org/10.1007/s11105-021-

506

01280-z

507

Kawasaki S, Miyake C, Kohchi T, Fujii S, Uchida M, Yokota A (2000) Responses of wild

508

watermelon to drought stress: accumulation of an ArgE homologue and citrulline in leaves

509

during water deficits. Plant Cell Physiol 41:864–873. https://doi.org/10.1093/pcp/pcd005

510

Lee DK, Redillas M, Jung H, Choi S, Kim YS, Kim JK (2018) A nitrogen molecular sensing

511

system, comprised of the ALLANTOINASE and UREIDE PERMEASE 1 genes, can be used

512

to monitor N status in rice. Front Plant Sci 9:444. https://doi.org/10.3389/fpls.2018.00444

513

Lescano I, Bogino MF, Martini C, Tessi TM, González CA, Schumacher K, Desimone M. (2020a)

514

Ureide permease 5 (atups5) connects cell compartments involved in ureide metabolism. Plant

515

Physiol, 182:1310–1325. https://doi.org/10.1104/pp.19.01136

516

Lescano CI, Martini C, González CA, Desimone M (2016) Allantoin accumulation mediated by

517

allantoinase downregulation and transport by Ureide Permease 5 confers salt stress tolerance

518

to Arabidopsis plants. Plant Mol Biol 91:581–595. https://doi.org/10.1007/s11103-016-0490-7

519

Lescano I, Devegili AM, Martini C, Tessi TM, González CA, Desimone M (2020b) Ureide

520

metabolism in Arabidopsis thaliana is modulated by C:N balance. J Plant Res 133:739–749.

521

https://doi.org/10.1007/s10265-020-01215-x

522

523

524

Ludwig RA (1993) Arabidopsis chloroplasts dissimilate L-arginine and L-citrulline for use as N

source. Plant Physiol 101:429–434. https://doi.org/10.1104/pp.101.2.429

Mikkelsen RL (1990) Biuret in urea fertilizer. Fertilizer Res 26: 311-318

525

526

527

Moore S, Stein WH (1954). A modified ninhydrin reagent for the photometric determination of

amino acids and related compounds. J Biol Chem, 211:907–913.

Nourimand M, Todd CD (2016) Allantoin increases cadmium tolerance in Arabidopsis via

528

activation

529

https://doi.org/10.1093/pcp/pcw162

antioxidant

mechanisms.

Plant

Cell

Physiol

57:2485–2496.

530

Ochiai K, Uesugi A, Masuda Y, Nishii M, Matoh T (2020) Overexpression of exogenous biuret

531

hydrolase in rice plants confers tolerance to biuret toxicity. Plant Direct 4:e00290.

532

https://doi.org/10.1002/pld3.290

533

Ogasawara S, Ezaki M, Ishida R, Sueyoshi K, Saito S, Hiradate Y, Kudo T, Obara M, Kojima S,

534

Uozumi N, Tanemura K, Hayakawa T (2021) Rice amino acid transporter-like 6 (OsATL6) is

535

involved in amino acid homeostasis by modulating the vacuolar storage of glutamine in roots.

536

Plant J 107:1616–1630. https://doi.org/10.1111/tpj.15403

537

538

Ogata T, Yamamoto M (1959) Effects of biuret on the metabolism of germinating plant. I. Jpn J

Soil Sci Plant Nutr 29:549-555 (in Japanese)

539

Pang Z, Chong J, Zhou G, de Lima Morais DA, Chang L, Barrette M, Gauthier C, Jacques PÉ, Li

540

S, Xia J (2021) MetaboAnalyst 5.0: narrowing the gap between raw spectra and functional

541

insights. Nucleic Acids Res 49:W388–W396. https://doi.org/10.1093/nar/gkab382

542

Redillas M, Bang SW, Lee DK, Kim YS, Jung H, Chung PJ, Suh JW, Kim JK (2019) Allantoin

543

accumulation through overexpression of ureide permease 1 improves rice growth under limited

544

nitrogen conditions. Plant Biotechnol J 17:1289–1301. https://doi.org/10.1111/pbi.13054

545

Robinson SL, Badalamenti JP, Dodge AG, Tassoulas LJ, Wackett LP (2018) Microbial

546

biodegradation of biuret: defining biuret hydrolases within the isochorismatase superfamily.

547

Environ microbiol 20:2099–2111. https://doi.org/10.1111/1462-2920.14094

548

Rocha PS, Sheikh M, Melchiorre R, Fagard M, Boutet S, Loach R, Moffatt B, Wagner C,

549

Vaucheret H, Furner I (2005) The Arabidopsis HOMOLOGY-DEPENDENT GENE

550

SILENCING1 gene codes for an S-adenosyl-L-homocysteine hydrolase required for DNA

551

methylation-dependent

552

https://doi.org/10.1105/tpc.104.028332

gene

silencing.

Plant

Cell

17:404–417.

553

Sakurai N, Ara T, Enomoto M, Motegi T, Morishita Y, Kurabayashi A, Iijima Y, Ogata Y,

554

Nakajima D, Suzuki H, Shibata D (2014) Tools and databases of the KOMICS web portal for

555

preprocessing, mining, and dissemination of metabolomics data. BioMed Res Int 2014:194812.

556

https://doi.org/10.1155/2014/194812

557

Schubert KR (1986) Products of biological nitrogen fixation in higher plants: Synthesis, transport,

558

and

559

https://doi.org/10.1146/annurev.pp.37.060186.002543

metabolism.

Annu

Rev

Plant

Physiol

37:539–574.

560

Soltabayeva A, Srivastava S, Kurmanbayeva A, Bekturova A, Fluhr R, Sagi M (2018) Early

561

senescence in older leaves of low nitrate-grown atxdh1 uncovers a role for purine catabolism

562

in n supply. Plant Physiol 178:1027–1044. https://doi.org/10.1104/pp.18.00795

563

Song Q, Joshi M, DiPiazza J, Joshi V (2020). Functional relevance of citrulline in the vegetative

564

tissues

565

https://doi.org/10.3389/fpls.2020.00512

watermelon

during

abiotic

stresses.

Front

Plant

Sci,

11:512.

566

Watanabe S, Matsumoto M, Hakomori Y, Takagi H, Shimada H, Sakamoto A (2014) The purine

567

metabolite allantoin enhances abiotic stress tolerance through synergistic activation of abscisic

568

acid metabolism. Plant Cell Environ 37:1022–1036. https://doi.org/10.1111/pce.12218

569

570

Webster GC, Berner RA, Gansa AN (1957) The effect of biuret on protein synthesis in plants.

Plant Physiol 32:60–61. https://doi.org/10.1104/pp.32.1.60

571

572

573

574

Yamaji N, Ma JF (2009) A transporter at the node responsible for intervascular transfer of silicon

in rice. Plant Cell 21:2878–2883. https://doi.org/10.1105/tpc.109.069831

Young EZ, Conway CF (1942) On the estimation of allantoin by the riminischryver reaction. J

Biol Chem 142:839-853. https://doi.org/10.1016/S0021-9258(18)45082-X

575

576

Statements and Declarations

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Funding: This work was supported in part by JSPS KAKENHI Grant Number JP19K05755.

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Competing interests: The authors have no relevant financial or non-financial interests to

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disclose.

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Author contributions: Kumiko Ochiai and Toru Matoh conceived and designed the research.

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Kumiko Ochiai and Yosuke Nomura performed experiments and analyzed the data. Asuka

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Uesugi generated biuret hydrolase-overexpressing rice lines. Kumiko Ochiai wrote the

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manuscript with input from other authors.

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Data availability: the data supporting the findings of this study are available within the article

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and its supplementary materials.

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Figure legend

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Fig. 1 Effects of biuret on dry weight (a, d), biuret concentration (b, e), and allantoin concentration

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(c, f) in roots and shoots of 7-day-old wild-type rice plants (a–c) and 9-day-old biuret hydrolase-

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overexpressing rice lines (d–f). Bars and circles represent mean and each sample, respectively.

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nd means not detected. (a–c) Wild-type plants were grown with 0, 0,1, and 0.3 mmol L-1 biuret

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supplemented in the culture solution. Ten seedlings were combined for a single sample. Different

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alphabets indicate significant difference among treatments in each organ (p < 0.05, Tukey’s test,

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n = 3). (d–f) Wild-type and two independent transgenic lines (B3-9-1 and B2-3-3) were grown

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with or without 0.3 mmol L-1 biuret in the culture solution. Four to six plants were combined into

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one sample. Gray and black bars represent control and biuret-treated plants, respectively.

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Asterisks indicate significant difference between the treatment (*p < 0.05; **p < 0.01, Welch’s t-

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test, n = 3). Different alphabets indicate significant difference in each organ (p < 0.05, Tukey’s

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test, n = 3).

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Fig. 2 Inhibitory effect of biuret for allantoinase activity. Crude extracts were prepared from

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shoots of 9-day-old rice seedlings hydroponically grown without biuret. Extracts were incubated

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at 30ºC with 10 mmol L-1 allantoin, 50 mmol L-1 Tricine-NaOH (pH8.0), 2mmol L-1 MnSO4, and

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the desired concentration of biuret for 30min. The amount of allantoic acid produced from

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allantoin was colorimetorically determined. Same shape symbols indicate a same crude extract.

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Crossbars represent the mean value. The means were not significantly different among treatments

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at 5% level (One-way ANOVA with blocking, n = 4).

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Fig. 3 Relative expression of genes related to purine degradation and ureide metabolisms in roots

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and shoots of 4 to 7-day old rice seedlings. Rice plants were hydroponically grown under the

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control condition and 0.3 mmol L-1 biuret toxicity. Data obtained from two independent trials,

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each with three replicates, are combined and shown. The relative expression levels of OsXO (a),

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OsUO (b), OsALNS (c), OsALN (d), OsAAH (e), OsUGlyAH (f), OsUAH (g), OsUPS1 (h),

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OsUPS2 (i), and OsUPS3 (j). The expression levels were normalized to the expression of

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Ubiquitin and Actin1 and expressed in log2 scale. Gray and black symbols indicate control and

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biuret treated samples, respectively. Crossbars indicate means of the six samples. Asterisks

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indicate statistically significant difference between the treatments at the time point. *p < 0.05;

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**p < 0.01; ***p < 0.001 (n = 6, Welch’s t-test). Numerical values above asterisks indicate log2

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fold-change relative to the control plants.

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Fig. 4 Principal component analysis of metabolomics profile of control and biuret-treated rice

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suspension cells. Rice cells were transferred into a medium without biuret or with 0.3 mmol L-1

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biuret and harvested 3 and 5 days after transfer. Closed symbols indicate control cells, and open

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symbols indicate biuret-treated cells. Circles indicate day 3 samples, and triangles indicate day 5

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samples.

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Fig. 5 Normalized peak intensities of differentially accumulated metabolites between control and

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biuret treated rice suspension cells. Peaks with significantly different intensity between the

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control-group and biuret-group are shown in the list (p < 0.05, Welch’s t-test, n = 4). RT column

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indicate retention time in second. In the formula column, the formula is shown when the formula

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is uniquely determined from the m/z value, blank when there are multiple possible candidates,

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and unknown when there are no candidates. D3C: day 3 control cell sample; D3B: day 3 biuret-

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treated cell sample; D5C: day 5 control cell sample; D5B: day 5 biuret-treated cell sample.

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Fig. 6 Free amino acids concentration (a) and allantoin concentration (b) in 8-day-old seedlings.

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Rice plants were hydroponically grown under the control condition and 0.3 mmol L -1 biuret

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toxicity. Five plants were combined for a single sample. Boxes indicate the mean of three samples,

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and symbols indicate each sample. Asterisks indicate a statistically significant difference between

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the treatments (Welch’s t-test). **p < 0.01; ***p < 0.001.

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Supplemental Materials

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Supplemental Fig. S1 Allantoin concentration in 9-day-old rice shoots measured by colorimetric

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and HPLC-UV. Plants were grown hydroponically under the three biuret treatments. Control:

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plants did not receive biuret; NB: plants were grown without biuret for three days after sowing,

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and with 0.3 mmol L-1 biuret supplemented in the culture solution from the fourth day; BN: plants

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were grown with 0.3 mmol L-1 biuret for 6 days after sowing and transferred to new culture

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solution without biuret on the seventh day. Fresh shoots of 9-day-old seedlings were ground under

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liquid N2 and extracted with 10-fold volume of distilled water. After centrifugation, the

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supernatant was used for allantoin determination. Gray boxes indicate mean allantoin

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concentration determined colorimetrically and black boxes indicate that determined by the HPLC

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method. Symbols indicate each sample. The statistical significance of the differences between the

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methods was determined through paired t-test (n = 2). ns: not significant.

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Supplemental Fig. S2 Pictures of 9-day-old wild-type rice seedlings and two biuret hydrolase-

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overexpressing lines. From left to right: wild type and overexpressing lines B3-9-1 and B2-3-3.

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Upper: seedlings grown in the control culture solution. Lower: seedlings grown in the culture

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solution supplemented with 0.3 mmol L-1 biuret. Bars show 10 cm.

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Supplemental Fig. S3 Relative expression levels of (a) OsALN and (b) OsUO in 3 to 9-day-old

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rice shoots in the preliminary experiment. Rice plants were hydroponically grown with or

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without 0.3 mmol L-1 biuret supplied in the culture solution. Measurements were made using

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plants grown in an independent trial from those shown in Figure 3. The relative expression

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levels were normalized to those of Ubiquitin and Actin1 and expressed on a log2 scale. Gray

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and black squares indicate control and biuret-treated plants, respectively; data represent the

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mean ± SD (n = 4). Asterisks indicate statistically significant differences between treatments at

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the time point (Welch’s t-test). *p < 0.05; **p < 0.01; ***p < 0.001. Numerical values above

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asterisks indicate log2 fold-change relative to the control plants.

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Supplemental Data S1 Peak intensities in Metabolome analysis.

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Fig. 1 Effects of biuret on dry weight (a, d), biuret concentration (b, e), and allantoin concentration

(c, f) in roots and shoots of 7-day-old wild-type rice plants (a–c) and 9-day-old biuret hydrolaseoverexpressing rice lines (d–f). Bars and circles represent mean and each sample, respectively.

nd means not detected. (a–c) Wild-type plants were grown with 0, 0,1, and 0.3 mmol L-1 biuret

supplemented in the culture solution. Ten seedlings were combined for a single sample. Different

alphabets indicate significant difference among treatments in each organ (p < 0.05, Tukey’s test,

n = 3). (d–f) Wild-type and two independent transgenic lines (B3-9-1 and B2-3-3) were grown

with or without 0.3 mmol L-1 biuret in the culture solution. Four to six plants were combined into

one sample. Gray and black bars represent control and biuret-treated plants, respectively.

Asterisks indicate significant difference between the treatment (*p < 0.05; **p < 0.01, Welch’s ttest, n = 3). Different alphabets indicate significant difference in each organ (p < 0.05, Tukey’s

test, n = 3).

Fig. 2 Inhibitory effect of biuret for allantoinase activity. Crude extracts were prepared from

shoots of 9-day-old rice seedlings hydroponically grown without biuret. Extracts were incubated

at 30ºC with 10 mmol L-1 allantoin, 50 mmol L-1 Tricine-NaOH (pH8.0), 2mmol L-1 MnSO4,

and the desired concentration of biuret for 30min. The amount of allantoic acid produced from

allantoin was colorimetorically determined. Same shape symbols indicate a same crude extract.

Crossbars represent the mean value. The means were not significantly different among

treatments at 5% level (One-way ANOVA with blocking, n = 4).