Skip to main content
SpringerLink
Log in

Induction of H2O2 in Transgenic Rice Leads to Cell Death and Enhanced Resistance to Both Bacterial and Fungal Pathogens

  • Published:
Transgenic Research Aims and scope Submit manuscript

Abstract

Oxidative burst, mediated by hydrogen peroxide (H2O2), has been recognized as a key component of plant defense response during an incompatible interaction. To determine if elevated levels of H2O2 lead to cell death, activation of defense genes and enhanced resistance to diverse pathogens, transgenic rice plants expressing a fungal glucose oxidase gene (GOX) were generated using both constitutive and inducible expression systems. Constitutive or wound/pathogen-induced expression of GOX also allowed us to determine the effectiveness of these systems in conferring long lasting resistance to various pathogens. Both constitutive and wound/pathogen-induced expression of GOX lead to increases in the endogenous levels of H2O2, which in turn caused cell death. Elevated levels of H2O2 also activated the expression of several defense genes and these transgenic plants showed enhanced resistance to both bacterial and fungal pathogens. In comparison to inducible expression, constitutive expression of GOX resulted in 3–10-fold higher levels of the GOX transcript and the corresponding enzymatic activity. Such increased levels of GOX, which would result in elevated levels of H2O2, caused improper seed set and decreased seed viability in transgenic plants constitutively expressing GOX. Our results suggest that pathogen inducible expression of heterologous genes may be a practical and robust way of generating broad spectrum disease resistance.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Alvarez ME and Lamb C (1997) Oxidative burst-mediated defense responses in plant disease resistance. In: Scandalios JG (ed.), Oxidative Stress and the Molecular Biology of Antioxidant Defenses. (pp. 815–839) Cold Spring Harbor Laboratory, Plainview, NY.

    Google Scholar 

  • Alvarez ME, Pennell RI, Meijer PJ, Ishikawa A, Dixon RA and Lamb CJ (1998) Reactive oxygen intermediates mediate a systemic signal network in the establishment of plant immunity. Cell 92: 773–784.

    Google Scholar 

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

    Google Scholar 

  • Bowles RJ (1990) Defense-related proteins in higher plants. Annu. Rev. Biochem. 59: 873–907.

    Google Scholar 

  • Bradley DJ, Kjellbom P and Lamb CJ (1992) Elicitor-and woundinduced oxidative cross-linking of a proline-rich plant cell wall protein: a novel, rapid defense response. Cell 70: 21–30.

    Google Scholar 

  • Brennan T and Frenkel C (1977) Involvement of hydrogen peroxide in the regulation of senescence in pear. Plant Phys. 59: 411–416.

    Google Scholar 

  • Brisson LF, Tenhaken R and Lamb CJ (1994) Function of oxidative cross-linking of cell wall structural proteins in plant disease resistance. Plant Cell 6: 1703–1712.

    Google Scholar 

  • Cazale AC, Rouet-Mayer MA, Barbier-Brygoo H, Mathieu Y and Lauriere C (1998) Oxidative burst and hypoosmotic stress in tobacco cell suspensions. Plant Physiol. 116: 659–669.

    Google Scholar 

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

    Google Scholar 

  • Chen L, Zhang S, Beachy RN and Fauquet CM (1998) A protocol for consistent, large-scale production of fertile transgenic rice plants. Plant Cell Rep. 18: 25–31.

    Google Scholar 

  • Chen Z, Silva H and Klessig DF (1993) Active oxygen species in the induction of plant systemic acquired resistance by salicylic acid. Science 262: 1883–1886.

    Google Scholar 

  • Chittoor JM, Leach JE and White FF (1997) Differential induction of a peroxidase gene family during infection of rice by Xanthomonas oryzae pv. oryzae. Mol. Plant Microbe Interact. 10: 861–871.

    Google Scholar 

  • Church GM and Gilbert M (1984) Genomic sequencing. Proc. Natl. Acad. Sci. USA 81: 1991–1995.

    Google Scholar 

  • Curtis MD, Rae AL, Rusu AG, Harrison SJ and Manners JM (1997) A peroxidase gene promoter induced by phytopathogens and methyl jasmonate in transgenic plants. Mol. Plant Microbe Interact. 10: 326–338.

    Google Scholar 

  • Dangl JL, Dietrich RA and Richberg MH (1996) Death don't have no mercy: cell death programs in plant-microbe interactions. Plant Cell 8: 1793–1807.

    Google Scholar 

  • del Pozo O and Lam E (1998) Caspases and programmed cell death in the hypersensitive response of plants to pathogens. Curr. Biol. 8: 1129–1132.

    Google Scholar 

  • Dixon RA and Lamb CJ (1990) Molecular communication in interactions between plants and microbial pathogens. Annu. Rev. Plant Physiol. Plant Mol. Biol. 41: 339–367.

    Google Scholar 

  • Esnault R and Chibbar RN (1997) Peroxidase and plant defense. LABPV Newsletter 10: 22–26.

    Google Scholar 

  • Frederick KR, Tung J, Emerick RS, Masiarz FR, Chamberlain SH, Vasavada A et al. (1990) Glucose oxidase from Aspergillus niger. Cloning, gene sequence, secretion from Saccharomyces cerevisiae and kinetic analysis of a yeast-derived enzyme. Biol. Chem. 265: 3793–3802.

    Google Scholar 

  • Greenberg JT, Guo A, Klessig DF and Ausubel FM (1994) Programmed cell death in plants: a pathogen-triggered response activated coordinately with multiple defense functions. Cell 77: 551–563.

    Google Scholar 

  • Greenberg JT (1996) Programmed cell death: a way of life for plants. Proc. Natl. Acad. Sci. USA 93: 12094–12097.

    Google Scholar 

  • Jabs T, Dietrich RA and Dangl JL (1996) Initiation of runaway cell death in an Arabidopsis mutant by extracellular superoxide. Science 273: 1853–1856.

    Google Scholar 

  • Kawasaki T, Henmi K, Ono E, Hatakeyama S, Megumi I, Satoh H et al. (1999) The small GTP-binding protein Rac is a regulator of cell death in plants. Proc. Natl. Acad. Sci. USA 96: 10922–10926.

    Google Scholar 

  • Kazan K, Murray FR, Goulter KC, Llewellyn DJ and Manners JM (1998) Induction of cell death in transgenic plants expressing a fungal glucose oxidase. Mol. Plant Microbe Interact. 11: 555–562.

    Google Scholar 

  • Keller T, Damude HG, Werner D, Doerner P, Dixon RA and Lamb CJ (1998) A plant homolog of the neutrophil NADPH oxidase gp91phox subunit gene encodes a plasma membrane protein with Ca+2 binding motifs. Plant Cell 10: 255–266.

    Google Scholar 

  • Kinoshita T, Imamura T, Nagai H and Shimotohno K (1992) Quanti-fication of gene expression over a wide range by the polymerase chain reaction. Annu. Rev. Biochem. 206: 231–235.

    Google Scholar 

  • Kiraly Z, El-Zahaby H, Galal A, Abdou S and Adam A (1993) Effect of oxygen free radicals on plant pathogenic bacteria and fungi and on some plant diseases. In: Mozsik G, Emerit J, Feher J, Matkovics B and Vincze A (eds), Oxygen Free Radicals and Scavengers in the Natural Sciences. (pp.9–19) Akademiai Kiado, Budapest.

    Google Scholar 

  • Koch E and Slusarenko A (1990) Arabidopsis is susceptible to infection by a downy mildew fungus. Plant Cell 2: 437–445.

    Google Scholar 

  • Lamb CJ (1998) Prospects for engineering enhanced durable disease resistance in crops. In: Whterlow JC, Armstrong DG, Fowden L and Riley R (eds), Feeding a World Population of More than Eight Billion People: A Challenge to Science. (pp. 183–190) Oxford University Press in association with the Rank Prize Funds, New York.

    Google Scholar 

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

    Google Scholar 

  • Lee YH, Yoon IS, Suh SC and Ki HI (2002) Enhanced disease resistance in transgenic cabbage and tobacco expressing a glucose oxidase from Aspergillus niger. Plant Cell Rep. 20: 857–863.

    Google Scholar 

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

    Google Scholar 

  • Mehdy MC (1994) Active oxygen species in plant defense against pathogens. Plant Physiol. 105: 467–472.

    Google Scholar 

  • Meier BM, Shaw N and Slusarenko AJ (1993) Spatial and temporal accumulation of defense gene transcripts in bean (Phaseolus vulgaris) leaves in relation to bacteria-induced hypersensitive cell death. Mol. Plant Microbe Interact. 6: 453–466.

    Google Scholar 

  • Mittler R, Del Pozo O, Meisel L and Lam E (1997) Pathogeninduced programmed cell death in plants, a possible defense mechanism. Dev. Genet. 21: 279–289.

    Google Scholar 

  • Mittler R, Feng X and Cohen M (1998) Post-transcriptional suppression of cytosolic ascorbate peroxidase expression during pathogen-induced programmed cell death in tobacco. Plant Cell 10: 461–473.

    Google Scholar 

  • Murray FR, Llewellyn DJ, Peacock WJ and Dennis ES (1997) Isolation of the glucose oxidase gene from Talaromyces flavus and characterisation of its role in the biocontrol of Verticillium dahliae. Curr. Genet. 32: 367–375.

    Google Scholar 

  • Nishi R, Hashimoto H, Kidou S, Uchimiya H and Kato A (1993) Isolation and characterization of a rice cDNA which encodes a ubiquitin protein and a 52 amino acid extension protein. Plant Mol Biol. 22: 159–161.

    Google Scholar 

  • Olson PD and Varner JE (1993) Hydrogen peroxide and lignification. Plant J. 4: 887–892.

    Google Scholar 

  • Pugin A, Frachisse J-M, Tavernier E, Bligny R, Gout E, Douce R et al. (1997) Early events induced by the elicitor cryptogein in tobacco cells: involvement of a plasma membrane NADPH oxidase and activation of glycolysis and the pentose phosphate pathway. Plant Cell 9: 2077–2091.

    Google Scholar 

  • Ryals JA, Neuenschwander UH, Willits MG, Molina A, Steiner H-Y and Hunt MD (1996) Systemic acquired resistance. Plant Cell 8: 1809–1819.

    Google Scholar 

  • Sambrook J, Fritsch EF and Maniatis T (1989) Molecular Cloning: A Laboratory Manual, II. Cold Spring Harbor Laboratory Press, New York.

    Google Scholar 

  • Schweizer P, Buchala A, Dudler R and Metraux J-P (1998) Induced systemic resistance in wounded rice plants. Plant J. 14: 475–481.

    Google Scholar 

  • Shirasu K, Dixon RA and Lamb CJ (1996) Signal transduction in plant immunity. Curr. Opin. Immunol. 8: 3–7.

    Google Scholar 

  • Shirasu K, Nakajima H, Rajasekhar VK, Dixon RA and Lamb CJ (1997) Salicylic acid potentiates an agonist-dependent gain control that amplifies pathogen signals in the activation of defense mechanisms. Plant Cell 9: 261–270.

    Google Scholar 

  • Song W-Y, Wang G-L, Chen L-L, Kim HS, Pi L-Y, Holsten T et al. (1995) A receptor kinase-like protein encoded by the rice disease resistance gene, Xa21. Science 270: 1804–1806.

    Google Scholar 

  • Takahashi H, Chen Z, Du H, Liu Y and Klessig DF (1997) Development of necrosis and activation of disease resistance in transgenic tobacco plants with severely reduced catalase levels. Plant J. 11: 993–1005.

    Google Scholar 

  • Tenhaken R, Levine A, Brisson LF, Dixon RA and Lamb CJ (1995) Function of the oxidative burst in hypersensitive disease resistance. Proc. Natl. Acad. USA 92: 4158–4163.

    Google Scholar 

  • Tiedemann AV (1997) Evidence for a primary role of active oxygen species in induction of host cell death during infection of bean leaves with Botrytis cinerea. Physiol. Mol. Plant Pathol. 50: 151–166.

    Google Scholar 

  • Tzeng DD and De Vay JE (1993) Role of oxygen radicals in plant disease development. Adv. Plant Pathol. 10: 1–34.

    Google Scholar 

  • Valent B, Farrall L and Chumley FG (1991) Magnaporthe grisea genes for pathogenicity and virulence identified through a series of backcrosses. Genetics 127: 87–101.

    Google Scholar 

  • Ward ER, Uknes SJ, Williams SC, Dincher SS, Weiderhold DL, Alexander DD et al. (1991) Coordinate gene activity in response to agents that induce systemic acquired resistance. Plant Cell 3: 1085–1094.

    Google Scholar 

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

    Google Scholar 

  • Wu G, Shortt BJ, Lawrence EB, Levine EB, Fitzsimmons KC and Shah DM (1995) Disease resistance conferred by expression of a gene encoding H2O2-generating glucose oxidase in transgenic potato plants. Plant Cell 7: 1357–1368.

    Google Scholar 

  • Wu G, Shortt BJ, Lawrence EB, Len J, Fitzsimmons KC, Levine EB, Raskin I et al. (1997) Activation of host defense mechanisms by elevated production of H2O2 in transgenic plants. Plant Physiol. 115: 427–435.

    Google Scholar 

  • Zhu Q and Lamb CJ (1991) Isolation and characterization of a rice gene encoding a basic chitinase. Mol. Gen. Genet. 226: 289–296.

    Google Scholar 

  • Zhu Q, Dabi T, Beeche A, Yamamoto R, Lawton MA and Lamb CJ (1995) Cloning and properties of a rice gene encoding phenylalanine ammonia-lyase. Plant Mol. Biol. 29: 535–550.

    Google Scholar 

Download references

Author information

Author notes

  1. Aardra Kachroo

    Present address: Department of Plant Pathology, University of Kentucky, Lexington, KY, 40546

  2. Qun Zhu

    Present address: Biotechnology Institute, Zhejiang University, Hangzhou, 310029, China

  3. Chris Lamb

    Present address: , Colney, Norwich, NR4 7UH, UK

Authors and Affiliations

  1. Department of Microbiology and Biotechnology Centre, M. S. University of Baroda, Baroda, 390002, India

    Aardra Kachroo, Rajesh Patkar, Jingping Zhong & Pamela Ronald

  2. Plant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA, 92037, USA

    Aardra Kachroo, Zuhua He & Qun Zhu

  3. Department of Plant Pathology, University of Kentucky, Lexington, KY, 40546

    Zuhua He & Bharat B. Chattoo

  4. Biotechnology Institute, Zhejiang University, Hangzhou, 310029, China

    Zuhua He, Debao Li & Chris Lamb

Authors
  1. Aardra Kachroo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Zuhua He
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rajesh Patkar
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Qun Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jingping Zhong
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Debao Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Pamela Ronald
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Chris Lamb
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Bharat B. Chattoo
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kachroo, A., He, Z., Patkar, R. et al. Induction of H2O2 in Transgenic Rice Leads to Cell Death and Enhanced Resistance to Both Bacterial and Fungal Pathogens. Transgenic Res 12, 577–586 (2003). https://doi.org/10.1023/A:1025896513472

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1025896513472

Search

Navigation