Skip to main content
SpringerLink
Log in

Activation of members of a MAPK module in β-glucan elicitor-mediated non-host resistance of soybean

  • Original Article
  • Published:
Planta Aims and scope Submit manuscript

Abstract

Plants recognize microbial pathogens by discriminating pathogen-associated molecular patterns from self-structures. We study the non-host disease resistance of soybean (Glycine max L.) to the oomycete, Phytophthora sojae. Soybean senses a specific molecular pattern consisting of a branched heptaglucoside that is present in the oomycetal cell walls. Recognition of this elicitor may be achieved through a β-glucan-binding protein, which forms part of a proposed receptor complex. Subsequently, soybean mounts a complex defense response, which includes the increase of the cytosolic calcium concentration, the production of reactive oxygen species, and the activation of genes responsible for the synthesis of phytoalexins. We now report the identification of two mitogen-activated protein kinases (MAPKs) and one MAPK kinase (MAPKK) that may function as signaling elements in triggering the resistance response. The use of specific antisera enabled the identification of GmMPKs 3 and 6 whose activity is enhanced within the signaling pathway leading to defense reactions. Elicitor specificity of MAPK activation as well as the sensitivity against inhibitors suggested these kinases as part of the β-glucan signal transduction pathway. An upstream GmMKK1 was identified based on sequence similarity to other plant MAPKKs and its interaction with the MAPKs was analyzed. Recombinant GmMKK1 interacted predominantly with GmMPK6, with concomitant phosphorylation of the MAPK protein. Moreover, a preferential physical interaction between GmMKK1 and GmMPK6 was demonstrated in yeast. These results suggest a role of a MAPK cascade in mediating β-glucan signal transduction in soybean, similar to other triggers that activate MAPKs during innate immune responses in plants.

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

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

Abbreviations

A9C:

Anthracene-9-carboxylate

DP:

Degree of polymerization

DPI:

Diphenyleneiodonium

EGTA:

Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid

EST:

Expressed sequence tag

IMAC:

Immobilized metal chelate affinity chromatography

MAPK:

(MPK) mitogen-activated protein kinase

MAPKK:

(MKK) MAPK kinase

MBP:

Myelin basic protein

NPPB:

5-Nitro-2-(3-phenylpropylamino)-benzoate

PAGE:

Polyacrylamide gelelectrophoresis

SA:

Salicylic acid

SSC:

Standard saline citrate

References

  • Asai T, Tena G, Plotnikova J, Willmann MR, Chiu W-L, Gomez-Gomez L, Boller T, Ausubel FM, Sheen J (2002) MAP kinase signalling cascade in Arabidopsis innate immunity. Nature 415:977–983

    Article  PubMed  CAS  Google Scholar 

  • Bögre L, Ligterink W, Meskiene I, Barker PJ, Heberle-Bors E, Huskisson NS, Hirt H (1997) Wounding induces the rapid and transient activation of a specific MAP kinase pathway. Plant Cell 9:75–83

    Article  PubMed  Google Scholar 

  • Boyle WJ, van der Geer P, Hunter T (1991) Phosphopeptide mapping and phosphoamino acid analysis by two-dimensional separation on thin-layer cellulose plates. Methods Enzymol 201:110–149

    Article  PubMed  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 282:152–160

    Google Scholar 

  • Brunner F, Rosahl S, Lee J, Rudd JJ, Geiler C, Kauppinen S, Rasmussen G, Scheel D, Nürnberger T (2002) Pep-13, a plant defense-inducing pathogen-associated pattern from Phytophthora transglutaminases. EMBO J 21:6681–6688

    Article  PubMed  CAS  Google Scholar 

  • Cardinale F, Jonak C, Ligterink W, Niehaus K, Boller T, Hirt H (2000) Differential activation of four specific MAPK pathways by distinct elicitors. J Biol Chem 275:36734–36740

    Article  PubMed  CAS  Google Scholar 

  • Cardinale F, Meskiene I, Ouaked F, Hirt H (2002) Convergence and divergence of stress-induced mitogen-activated protein kinase signaling pathways at the level of two distinct mitogen-activated protein kinase kinases. Plant Cell 14:703–711

    PubMed  CAS  Google Scholar 

  • Chinchilla D, Bauer Z, Regenass M, Boller T, Felix G (2006) The Arabidopsis receptor kinase FLS2 binds flg22 and determines the specificity of flagellin perception. Plant Cell 18:465–476

    Article  PubMed  CAS  Google Scholar 

  • Contor L, Lamy F, Lecocq RE (1987) Use of electroblotting to detect and analyze phosphotyrosine containing peptides separated by two-dimensional gel electrophoresis. Anal Biochem 160:414–420

    Article  PubMed  CAS  Google Scholar 

  • Corpet F (1988) Multiple sequence alignment with hierarchical clustering. Nucl Acids Res 16:10881–10890

    Article  PubMed  CAS  Google Scholar 

  • Cosio EG, Frey T, Verduyn R, van Boom J, Ebel J (1990) High-affinity binding of a synthetic heptaglucoside and fungal glucan phytoalexin elicitors to soybean membranes. FEBS Lett 271:223–226

    Article  PubMed  CAS  Google Scholar 

  • del Pozo O, Pedley KF, Martin GB (2004) MAPKKKα is a positive regulator of cell death associated with both plant immunity and disease. EMBO J 23:3072–3082

    Article  PubMed  CAS  Google Scholar 

  • Ebel J, Mithöfer A (1998) Early events in the elicitation of plant defense. Planta 206:335–348

    Article  CAS  Google Scholar 

  • Felix G, Duran JD, Volko S, Boller T (1999) Plants have a sensitive perception system for the most conserved domain of bacterial flagellin. Plant J 18:265–276

    Article  PubMed  CAS  Google Scholar 

  • Fliegmann J, Mithöfer A, Wanner G, Ebel J (2004) An ancient enzyme domain hidden in the putative β-glucan elicitor receptor of soybean may play an active part in the perception of pathogen-associated molecular patterns during broad host resistance. J Biol Chem 279:1132–1140

    Article  PubMed  CAS  Google Scholar 

  • Fliegmann J, Schüler G, Boland W, Ebel J, Mithöfer A (2003) The role of octadecanoids and functional mimics in soybean defense responses. Biol Chem 384:437–446

    Article  PubMed  CAS  Google Scholar 

  • Hall TA (1999) BioEdit. A user-friendly biological sequence alignment and analysis program for Windows 95/98/NT. Nucl Acid Symp Ser 41:95–98

    CAS  Google Scholar 

  • He P, Shan L, Lin N-C, Martin GB, Kemmerling B, Nürnberger T, Sheen J (2006) Specific bacterial suppressors of MAMP signaling upstream of MAPKKK in Arabidopsis innate immunity. Cell 125:563–575

    Article  PubMed  CAS  Google Scholar 

  • Hille A, Purwin C, Ebel J (1982) Induction of enzymes of phytoalexin synthesis in cultured soybean cells by an elicitor from Phytophthora megasperma f. sp. glycinea. Plant Cell Rep 1:123–127

    Article  CAS  Google Scholar 

  • Huang Y, Li H, Gupta R, Morris PC, Luan S, Kieber JJ (2000) AtMPK4, an Arabidopsis homolog of mitogen-activated protein kinase, is activated in vitro by AtMEK1 through threonine phosphorylation. Plant Physiol 122:1301–1310

    Article  PubMed  CAS  Google Scholar 

  • Janeway CA Jr, Medzhitov R (2002) Innate immune recognition. Annu Rev Immunol 20:197–216

    Article  PubMed  CAS  Google Scholar 

  • Jonak C, Kiegerl S, Ligterink W, Barker PJ, Huskisson NS, Hirt H (1996) Stress signaling in plants: a mitogen-activated protein kinase pathway is activated by cold and drought. Proc Natl Acad Sci USA 93:11274–11279

    Article  PubMed  CAS  Google Scholar 

  • Jonak C, Pay A, Bögre L, Hirt H, Heberle-Bors E (1993) The plant homologue of MAP kinase is expressed in a cell cycle-dependent and organ-specific manner. Plant J 3:611–617

    Article  PubMed  CAS  Google Scholar 

  • Kiegerl S, Cardinale F, Siligan C, Gross A, Baudouin E, Liwosz A, Eklöf S, Till S, Bögre L, Hirt H, Meskiene I (2000) SIMKK, a mitogen-activated protein kinase (MAPK) kinase, is a specific activator of the salt stress-induced MAPK, SIMK. Plant Cell 12:2247–2258

    Article  PubMed  CAS  Google Scholar 

  • Kroj T, Rudd JJ, Nürnberger T, Gäbler Y, Lee J, Scheel D (2003) Mitogen-activated protein kinases play an essential role in oxidative burst-independent expression of pathogenesis-related genes in parsley. J Biol Chem 278:2256–2264

    Article  PubMed  CAS  Google Scholar 

  • Lebrun-Garcia A, Ouaked F, Chiltz A, Pugin A (1998) Activation of MAPK homologues by elicitors in tobacco cells. Plant J 15:773–781

    Article  PubMed  CAS  Google Scholar 

  • Lee J, Klessig DF, Nürnberger T (2001b) A harpin binding site in tobacco plasma membranes mediates activation of the pathogenesis-related gene HIN1 independent of extracellular calcium but dependent on mitogen-activated protein kinase activity. Plant Cell 13:1079–1093

    Article  CAS  Google Scholar 

  • Lee J, Rudd JJ, Macioszek VK, Scheel D (2004) Dynamic changes in the localization of MAPK cascade components controlling pathogenesis-related (PR) gene expression during innate immunity in parsley. J Biol Chem 279:22440–22448

    Article  PubMed  CAS  Google Scholar 

  • Lee S, Hirt H, Lee Y (2001a) Phosphatidic acid activates a wound-activated MAPK in Glycine max. Plant J 26:479–486

    Article  CAS  Google Scholar 

  • Liu YH, Zhang SQ, Klessig DF (2000) Molecular cloning and characterization of a tobacco MAP kinase kinase that interacts with SIPK. Mol Plant-Microbe Interact 13:118–124

    PubMed  CAS  Google Scholar 

  • MAPK Group (2002) Mitogen-activated protein kinase cascades in plants: a new nomenclature. Trends Plant Sci 7:301–308

    Article  Google Scholar 

  • Medzhitov R, Janeway CA Jr (1997) Innate immunity: the virtues of a nonclonal system of recognition. Cell 91:295–298

    Article  PubMed  CAS  Google Scholar 

  • Mithöfer A, Bhagwat AA, Feger M, Ebel J (1996) Suppression of fungal β-glucan-induced plant defence in soybean (Glycine max L.) by cyclic 1,3–1,6-β-glucans from the symbiont Bradyrhizobium japonicum. Planta 199:270–275

    Article  Google Scholar 

  • Mithöfer A, Daxberger A, Fromhold-Treu D, Ebel J (1997) Involvement of an NAD(P)H oxidase in the elicitor-inducible oxidative burst of soybean. Phytochemistry 45:1101–1107

    Article  Google Scholar 

  • Mithöfer A, Ebel J, Bhagwat AA, Boller T, Neuhaus-Url G (1999) Transgenic aequorin monitors cytosolic calcium transients in soybean cells challenged with β-glucan or chitin elicitors. Planta 207:566–574

    Article  Google Scholar 

  • Mithöfer A, Ebel J, Felle H (2005) Cation fluxes cause plasma membrane depolarization involved in β-glucan elicitor-signaling in soybean roots. Mol Plant-Microbe Interact 18:983–990

    PubMed  Google Scholar 

  • Mithöfer A, Fliegmann J, Daxberger A, Ebel C, Neuhaus U, Bhagwat AA, Keister DL, Ebel J (2001) Induction of H2O2 synthesis by β-glucan elicitors in soybean is independent on cytosolic calcium transients. FEBS Lett 508:191–195

    Article  PubMed  Google Scholar 

  • Mithöfer A, Fliegmann J, Neuhaus-Url G, Schwarz H, Ebel J (2000) The hepta-β-glucoside elicitor-binding proteins from legumes represent a putative receptor family. Biol Chem 381:705–713

    Article  PubMed  Google Scholar 

  • Mizoguchi T, Hayashida N, Yamaguchi-Shinozaki K, Kamada H, Shinozaki K (1993) AtMPKs: a gene family of plant MAP kinases in Arabidopsis thaliana. FEBS Lett 336:440–444

    Article  PubMed  CAS  Google Scholar 

  • Munnik T, Ligterink W, Meskiene I, Calderini O, Beyerly J, Musgrave A, Hirt H (1999) Distinct osmo-sensing protein kinase pathways are involved in signalling moderate and severe hyper-osmotic stress. Plant J 20:381–388

    Article  PubMed  CAS  Google Scholar 

  • Nakagami H, Pitzschke A, Hirt H (2005) Emerging MAPK pathways in plant stress signaling. Trends Plant Sci 10:339–346

    Article  PubMed  CAS  Google Scholar 

  • Nühse TS, Peck SC, Hirt H, Boller T (2000) Microbial elicitors induce activation and dual phosphorylation of the Arabidopsis thaliana MAPK6. J Biol Chem 275:7521–7526

    Article  PubMed  Google Scholar 

  • Nürnberger T, Scheel D (2001) Signal transmission in the plant immune response. Trends Plant Sci 6:372–379

    Article  PubMed  Google Scholar 

  • Nürnberger T, Brunner F, Kemmerling B, Piater L (2004) Innate immunity in plants and animals: similarities and obvious differences. Immunol Rev 198:249–266

    Article  PubMed  Google Scholar 

  • Pedley KF, Martin GB (2004) Identification of MAPKs and their possible MAPKK activators involved in the Pto-mediated defense response of tomato. J Biol Chem 279:49229–49235

    Article  PubMed  CAS  Google Scholar 

  • Pedley KF, Martin GB (2005) Role of mitogen-activated protein kinases in plant immunity. Curr Opin Plant Biol 8:541–547

    Article  PubMed  CAS  Google Scholar 

  • Ren D, Yang H, Zhang S (2002) Cell death mediated by MAPK is associated with hydrogen peroxide production in Arabidopsis. J Biol Chem 277:559–565

    Article  PubMed  CAS  Google Scholar 

  • Romeis T (2001) Protein kinases in the plant defence response. Curr Opin Plant Biol 4:407–414

    Article  PubMed  CAS  Google Scholar 

  • Seo S, Okamoto M, Seto H, Ishizuka K, Sano H, Ohashi Y (1995) Tobacco MAP kinase: a possible mediator in wound signal transduction pathways. Science 270:1988–1992

    Article  PubMed  CAS  Google Scholar 

  • Sharp JK, McNeil M, Albersheim P (1984b) The primary structures of one elicitor-active and seven elicitor-inactive hexa(β-D-glucopyranosyl)-D-glucitols isolated from the mycelial walls of Phytophthora megasperma f. sp. glycinea. J Biol Chem 259:11321–11336

    CAS  Google Scholar 

  • Sharp JK, Valent B, Albersheim P (1984a) Purification and partial characterization of a beta-glucan fragment that elicits phytoalexin accumulation in soybean. J Biol Chem 259:11312–11320

    CAS  Google Scholar 

  • Stäb MR, Ebel J (1987) Effects of Ca2+ on phytoalexin induction by fungal elicitor in soybean cells. Arch Biochem Biophys 257:416–423

    Article  PubMed  Google Scholar 

  • Stratmann JW, Ryan CA (1997) Myelin basic protein kinase activity in tomato leaves is induced systemically by wounding and increases in response to systemin and oligosaccharide elicitors. Proc Natl Acad Sci USA 94:11085–11089

    Article  PubMed  CAS  Google Scholar 

  • Suzuki K, Shinshi H (1995) Transient activation and tyrosine phosphorylation of a protein kinase in tobacco cells treated with a fungal elicitor. Plant Cell 7:639–647

    Article  PubMed  CAS  Google Scholar 

  • Taylor ATS, Kim J, Low PS (2001) Involvement of mitogen-activated protein kinase activation in the signal-transduction pathways of the soya bean oxidative burst. Biochem J 355:795–803

    PubMed  CAS  Google Scholar 

  • Tena G, Asai T, Chiu W-L, Sheen J (2001) Plant mitogen-activated protein kinase signaling cascades. Curr Opin Plant Biol 4:392–400

    Article  PubMed  CAS  Google Scholar 

  • Yang K-Y, Liu Y, Zhang S (2001) Activation of a mitogen-activated protein kinase pathway is involved in disease resistance in tobacco. Proc Natl Acad Sci USA 98:741–746

    Article  PubMed  CAS  Google Scholar 

  • Zhang S, Klessig DF (1997) Salicylic acid activates a 48-kD MAP kinase in tobacco. Plant Cell 9:809–824

    Article  PubMed  CAS  Google Scholar 

  • Zhang S, Klessig DF (2001) MAPK cascades in plant defense signaling. Trends Plant Sci 6:520–527

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgment

This work was supported by the Deutsche Forschungsgemeinschaft (SFB369).

Author information

Author notes

  1. Wilco Ligterink

    Present address: Laboratorium voor Plantenfysiologie, Botanisch Centrum, Arboretumlaan 4, 6703, Wageningen, The Netherlands

Authors and Affiliations

  1. Department Biologie I/Botanik, Ludwig-Maximilians-Universität, Menzinger Str. 67, 80638, München, Germany

    Andrea Daxberger, Andrea Nemak, Judith Fliegmann & Jürgen Ebel

  2. Max-Planck-Institut für Chemische Ökologie, Bioorganische Chemie, Hans-Knöll-Str. 8, 07745, Jena, Germany

    Axel Mithöfer

  3. Department für Pflanzenmolekularbiologie, Max F. Perutz Laboratories, University of Vienna, Dr. Bohrgasse 9, 1030, Vienna, Austria

    Heribert Hirt

Authors
  1. Andrea Daxberger
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Andrea Nemak
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Axel Mithöfer
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Judith Fliegmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wilco Ligterink
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Heribert Hirt
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jürgen Ebel
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jürgen Ebel.

Additional information

The nucleotide sequences encoding the MAPKs and MAPKK1 from soybean can be accessed through the GenBank database under GenBank accession numbers AF104247, AF329506, and AY070230.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Cite this article

Daxberger, A., Nemak, A., Mithöfer, A. et al. Activation of members of a MAPK module in β-glucan elicitor-mediated non-host resistance of soybean. Planta 225, 1559–1571 (2007). https://doi.org/10.1007/s00425-006-0442-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00425-006-0442-6

Keywords

Search

Navigation