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The cantharidin-induced oxidative burst in tobacco BY-2 cell suspension cultures

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Summary

The in vivo induction of H2O2 production was tested on tobacco cell suspension cultures (Nicotiana tabacum cv. Bright Yellow-2). The measurement of H2O2 was based on the oxidation of 3,5-dichloro-2-hydroxybenzensulfonic acid by endogenous peroxidases and spectrophotometric detection after reaction with 4-aminoanti-pyrine. The phosphatase inhibitor cantharidin induced a transient increase in H2O2 synthesis. The timing of the H2O2 production, the level of induction by cantharidin and the background H2O2 production were dependent on the tobacco cell concentration used. A concentration curve of cantharidin revealed saturating kinetics for the H2O2 detection (E50=46 to 70 μM, Emax=101 to 128 μmol/h·g fresh weight). An inhibitor study with the tobacco BY-2 cells showed high inhibitions of the H2O2 induction with the flavin analogues diphenylene iodonium (I50=1.26μM) and acridine orange and with membrane-permeative thiol reagents (N-ethyl maleimide, N-pyrene maleimide, iodoacetate); whereas the nonpermeative thiol reagentp-chloromercuribenzoic acid was ineffective. Therefore, the induction of H2O2 production with phosphatase inhibitors (cantharidin) showed comparable properties to the elicitor-induced oxidative-burst response in other plant cells.

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Abbreviations

AcOr:

acridine orange

AOS:

active-oxygen species

BY-2:

Bright Yellow-2

pCMBS:

p-chloromercuribenzoic acid

DHBS:

3,5-dichloro-2-hydroxybenzenesulfonic acid

DMSO:

dimethylsulfoxide

DPI:

diphenylene iodonium

EtOH:

ethanol

H2O2 :

hydrogen peroxide

HRP:

horseradish peroxidase

MS:

Murashige and Skoog

NEM:

N-ethyl maleimide

NPM:

N-pyrene maleimide

O −2 :

superoxide

SOD:

superoxide dismutase

References

  • Alan AC, Fluhr R (1997) Two distinct sources of elicited reactive oxygen species in tobacco epidermal cells. Plant Cell 9: 1559–1572

    Google Scholar 

  • Auh C-K, Murphy TM (1995) Plasma membrane redox enzyme is involved in the synthesis of O2 − and H2O2 byPhytophthora elicitor-stimulated rose cells. Plant Physiol 107: 1241–1247

    Google Scholar 

  • Baker CJ, Orlandi EW (1995) Active oxygen in plant pathogenesis. Annu Rev Phytopathol 33: 299–321

    Google Scholar 

  • Bolwell GP, Wojtaszek P (1997) Mechanisms for the generation of reactive oxygen species in plant defence: a broad perspective. Physiol Mol Plant Pathol 51: 347–366

    Google Scholar 

  • —, Butt VS, Davies DR, Zimmerlin A (1995) The origin of the oxidative burst in plants. Free Radie Res Commun 23: 517–532

    Google Scholar 

  • —, Davies DR, Gerrish C, Auh C-K, Murphy TM (1998) Comparative biochemistry of the oxidative burst produced by rose and French bean cells reveals two distinct mechanisms. Plant Physiol 116: 1379–1385

    Google Scholar 

  • Cervone F, Castoria R, Leckie F, De Lorenze G (1997) Perception of fungal elicitors and signal transduction. In: Aducci P (ed) Signal transduction in plants. Birkhäuser, Basel, pp 153–177

    Google Scholar 

  • Chamnongpol S, Willekens H, Moeder W, Langebartels C, Sandermann H, Van Montagu M, Inzé D, Van Camp W (1998) Defense activation and enhanced pathogen tolerance induced by H2O2 in transgenic tobacco. Proc Natl Acad Sci USA 95: 5818–5823

    Google Scholar 

  • Chandra S, Low PS (1995) Role of phosphorylation in elicitation of the oxidative burst in cultured soybean cells. Proc Natl Acad Sci USA 92: 4120–4123

    Google Scholar 

  • Dixon RA, Harrison MJ, Lamb CJ (1994) Early events in the activation of plant defense responses. Annu Rev Phytopathol 32: 479–501

    Google Scholar 

  • Doussiere J, Vignais PV (1992) Diphenylene iodonium as an inhibitor of the NADPH oxidase complex of bovine neutrophils: factors controlling the inhibitory potency of diphenylene iodonium in a cell-free system of oxidase activation. Eur J Biochem 208: 61–71

    Google Scholar 

  • Ebel J, Cosio EG (1994) Elicitors of plant defense responses. Int Rev Cytol 148: 1–36

    Google Scholar 

  • Felix G, Regenass M, Spanu P, Boiler T (1994) The protein phosphatase inhibitor calyculin A mimics elicitor action in plant cells and induces rapid hyperphosphorylation of specific proteins as revealed by pulse labelling with [32P] phosphate. Proc Natl Acad Sci USA 91: 952–956

    Google Scholar 

  • Hahn MG (1996) Microbial elicitors and their receptors in plants. Annu Rev Phytopathol 34: 387–412

    Google Scholar 

  • Hammond-Kosack KE, Jones JDG (1996) Resistance gene dependent plant defense responses. Plant Cell 8: 1773–1791

    Google Scholar 

  • Lamb C, Dixon RA (1997) The oxidative burst in plant disease resistance. Annu Rev Plant Physiol Plant Mol Biol 48: 251–275

    Google Scholar 

  • Legendre L, Rueter S, Heinstein PF, Low PS (1993) Characterization of the oligogalacturonide-induced oxidative burst in cultured soybean (Glycine max) cells. Plant Physiol 102: 233–240

    Google Scholar 

  • Levine A, Tenhaken R, Dixon R, Lamb C (1994) H2O2 from the oxidative burst orchestrates the plant hypersensitive disease resistance response. Cell 79: 583–593

    Google Scholar 

  • Low PS, Merida JR (1996) The oxidative burst in plant defense: function and signal transduction. Physiol Plant 96: 533–542

    Google Scholar 

  • Mathieu Y, Sanchez FJ, Droillard MJ, Lapous D, Laurrière C, Guern J (1996) Involvement of protein phosphorylation in the early steps of transduction of the oligogalacturonide signal in tobacco cells. Plant Physiol Biochem 34: 399–408

    Google Scholar 

  • Mehdy MC, Sharma YK, Sathasivan K, Bays NW (1996) The role of activated oxygen species in plant disease resistance. Physiol Plant 98: 365–374

    Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473–497

    Google Scholar 

  • Murphy TM, Auh C-K (1996) The superoxide synthases of plasma membrane preparations from cultured rose cells. Plant Physiol 110: 621–629

    Google Scholar 

  • —, Asard H, Cross AR (1998) Possible sources of reactive oxygen during the oxidative burst in plants. In: Asard H, Bérczi A, Caubergs R (eds) Plasma membrane redox systems and their role in biological stress and disease. Kluwer, Dordrecht (in press)

    Google Scholar 

  • Nagata T, Nemoto Y, Hasezawa S (1992) Tobacco BY-2 cell line as the “HeLa” cell in the cell biology of higher plants. Int Rev Cytol 132: 1–30

    Google Scholar 

  • Nürnberger T, Nennstiel D, Jabs T, Sacks WR, Hahlbrock K, Scheel D (1994) High affinity binding of a fungal oligopeptide elicitor to parsley plasma membranes trigger multiple defense responses. Cell 78: 449–460

    Google Scholar 

  • O'Donnell VB, Smith GC, Jones OT (1994) Involvement of phenyl radicals in iodonium inhibition of flavoenzymes. Mol Pharmacol 46: 778–785

    Google Scholar 

  • Pinton R, Cakmak I, Marschner H (1994) Zinc deficiency enhanced NAD(P)H-dependent superoxide radical production in plasma membrane vesicles isolated from roots of bean plants. J Exp Bot 45: 45–50

    Google Scholar 

  • Sutherland MW (1991) The generation of oxygen radicals during the plant responses to infection. Physiol Mol Plant Pathol 39: 79–83

    Google Scholar 

  • Svalheim O, Robertson B (1993) Elicitation of H2O2 production in cucumber hypocotyl segments by oligo-1,4-α-D-galacturonides and oligo-β-glucan preparations from cell walls ofPhytophthora megasperma f. sp.glycinea. Physiol Plant 88: 675–681

    Google Scholar 

  • Tenhaken R, Rubel C (1997) Cloning of soybean NADPH-oxidase subunits by expression screening. In: Abstract book of the International Conference on Plasma Membrane Redox Systems, Antwerp, April 1998, p 57

  • Van Gestelen P, Asard H, Caubergs RJ (1997) Solubilization and separation of a plant plasma membrane NADPH-O2 synthase from other NAD(P)H oxidoreductases. Plant Physiol 115: 543–550

    Google Scholar 

  • - - Horemans N, Caubergs RJ (1998) Superoxide producing NAD(P)H-oxidases in plasma membrane vesicles from elicitor responsive bean plants. Physiol Plant (in press)

  • Willekens H, Chamnongpol S, Davey M, Schraudner M, Langebartels C, Van Montagu M, Inzé D, Van Camp W (1997) Catalase is a sink for H2O2 and is indispensable for stress defense in C3 plants. EMBO J 16: 4806–4816

    Google Scholar 

  • Wojtaszek P (1997) Oxidative burst: an early plant response to pathogen infection. Biochem J 322: 681–692

    Google Scholar 

  • Xing T, Higgins VJ, Blumwald E (1997) Race-specific elicitors ofCladosporiumfulvum promote translocation of cytosolic components of NADPH oxidase to the plasma membrane of tomato cells. Plant Cell 9: 249–259

    Google Scholar 

  • Yahraus T, Chandra S, Legendre L, Low PS (1995) Evidence for a mechanically induced oxidative burst. Plant Physiol 109: 1259–1266

    Google Scholar 

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Authors and Affiliations

  1. Department of Biology, University of Antwerp, Groenenborgerlaan 171, B-2020, Antwerp, Belgium

    P. Van Gestelen, P. Ledeganck, I. Wynant, R. J. Caubergs & H. Asard

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  1. P. Van Gestelen
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  2. P. Ledeganck
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  3. I. Wynant
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  4. R. J. Caubergs
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  5. H. Asard
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Van Gestelen, P., Ledeganck, P., Wynant, I. et al. The cantharidin-induced oxidative burst in tobacco BY-2 cell suspension cultures. Protoplasma 205, 83–92 (1998). https://doi.org/10.1007/BF01279297

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  • DOI: https://doi.org/10.1007/BF01279297

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