Plant Cell Reports

, Volume 35, Issue 5, pp 1169–1185 | Cite as

Magnaporthe oryzae effectors MoHEG13 and MoHEG16 interfere with host infection and MoHEG13 counteracts cell death caused by Magnaporthe-NLPs in tobacco

  • Valerie Mogga
  • Rhoda Delventhal
  • Denise Weidenbach
  • Samantha Langer
  • Philipp M. Bertram
  • Karsten Andresen
  • Eckhard Thines
  • Thomas Kroj
  • Ulrich Schaffrath
Original Article

Abstract

Key message

Adapted pathogens are able to modulate cell responses of their hosts most likely due to the activity of secreted effector molecules thereby enabling colonisation by ostensible nonhost pathogens.

Abstract

It is postulated that host and nonhost pathogens of a given plant species differ in their repertoire of secreted effector molecules that are able to suppress plant resistance. We pursued the strategy of identifying novel effectors of Magnaporthe oryzae, the causal agent of blast disease, by comparing the infection process of closely related host vs. nonhost Magnaporthe species on barley (Hordeum vulgare L.). When both types of pathogen simultaneously attacked the same cell, the nonhost isolate became a successful pathogen possibly due to potent effectors secreted by the host isolate. Microarray studies led to a set of M. oryzae Hypothetical Effector Genes (MoHEGs) which were classified as Early- and LateMoHEGs according to the maximal transcript abundance during colonization of barley. Interestingly, orthologs of these MoHEGs from a nonhost pathogen were similarly regulated when investigated in a host situation, suggesting evolutionary conserved functions. Knockout mutants of MoHEG16 from the group of EarlyMoHEGs were less virulent on barley and microscopic studies revealed an attenuated transition from epidermal to mesophyll colonization. MoHEG13, a LateMoHEG, was shown to antagonize cell death induced by M. oryzae Necrosis-and ethylene-inducing-protein-1 (Nep1)-like proteins in Nicotiana benthamiana. MoHEG13 has a virulence function as a knockout mutant showed attenuated disease progression when inoculated on barley.

Keywords

Magnaporthe oryzae Barley Effector proteins Necrosis 

Supplementary material

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Supplementary material 1 (DOCX 18 kb)
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Supplementary material 2 (DOCX 15 kb)
299_2016_1943_MOESM3_ESM.pdf (219 kb)
Supplementary material 3 (PDF 218 kb)
299_2016_1943_MOESM4_ESM.pdf (295 kb)
Supplementary material 4 (PDF 295 kb)
299_2016_1943_MOESM5_ESM.pdf (204 kb)
Supplementary material 5 (PDF 203 kb)

References

  1. Bailey BA (1995) Purification of a protein from culture filtrates of Fusarium oxysporum that induces ethylene and necrosis in leaves of Erythroxylum coca. Phytopathology 85:1250–1255CrossRefGoogle Scholar
  2. Benjamini Y, Hochberg Y (1995) Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc Ser B (Methodol) 57:289–300Google Scholar
  3. Catlett NL, Lee DN, Yoder OC, Turgeon BG (2003) Split-marker recombination for efficient targeted deletion of fungal genes. Fungal Genet Newsl 50:9–11Google Scholar
  4. Cesari S, Thilliez G, Ribot C, Chalvon V, Michel C, Jauneau A, Rivas S, Alaux L, Kanzaki H, Okuyama Y, Morel J-B, Fournier E, Tharreau D, Terauchi R, Kroj T (2013) The rice resistance protein pair RGA4/RGA5 recognizes the Magnaporthe oryzae effectors AVR-Pia and AVR1-CO39 by direct binding. Plant Cell 25:1463–1481CrossRefPubMedPubMedCentralGoogle Scholar
  5. Chen S, Songkumarn P, Venu RC, Gowda M, Bellizzi M, Hu J, Liu W, Ebbole D, Meyers B, Mitchell T, Wang G-L (2013) Identification and characterization of in planta-expressed secreted effector proteins from Magnaporthe oryzae that induce cell death in rice. Mol Plant Microbe Interact 26:191–202CrossRefPubMedGoogle Scholar
  6. Couch BC, Kohn LM (2002) A multilocus gene genealogy concordant with host preference indicates segregation of a new species, Magnaporthe oryzae, from M. grisea. Mycologia 94:683–693CrossRefPubMedGoogle Scholar
  7. Dean RA, Talbot NJ, Ebbole DJ, Farman ML, Mitchell TK, Orbach MJ, Thon M, Kulkarni R, Xu J-R, Pan H, Read ND, Lee Y-H, Carbone I, Brown D, Oh YY, Donofrio N, Jeong JS, Soanes DM, Djonovic S, Kolomiets E, Rehmeyer C, Li W, Harding M, Kim S, Lebrun M-H, Bohnert H, Coughlan S, Butler J, Calvo S, Ma L-J, Nicol R, Purcell S, Nusbaum C, Galagan JE, Birren BW (2005) The genome sequence of the rice blast fungus Magnaporthe grisea. Nature 434:980–986CrossRefPubMedGoogle Scholar
  8. Delventhal R, Falter C, Strugala R, Zellerhoff N, Schaffrath U (2014) Ectoparasitic growth of Magnaporthe on barley triggers expression of the putative barley wax biosynthesis gene CYP96B22 which is involved in penetration resistance. BMC Plant Biol 14:26CrossRefPubMedPubMedCentralGoogle Scholar
  9. Dudler R, Hertig C (1992) Structure of an mdr-like gene from Arabidopsis thaliana. Evolutionary implications. J Biol Chem 267:5882–5888PubMedGoogle Scholar
  10. Eckert M, Maguire K, Urban M, Foster S, Fitt B, Lucas J, Hammond-Kosack K (2005) Agrobacterium tumefaciens-mediated transformation of Leptosphaeria spp. and Oculimacula spp. with the reef coral gene DsRed and the jellyfish gene gfp. FEMS Microbiol Lett 253:67–74CrossRefPubMedGoogle Scholar
  11. Faivre-Rampant O, Thomas J, Allegre M, Morel JB, Tharreau D, Notteghem JL, Lebrun MH, Schaffrath U, Piffanelli P (2008) Characterization of the model system rice-Magnaporthe for the study of nonhost resistance in cereals. New Phytol 180:899–910CrossRefPubMedGoogle Scholar
  12. Fisher MC, Henk DA, Briggs CJ, Brownstein JS, Madoff LC, McCraw SL, Gurr SJ (2012) Emerging fungal threats to animal, plant and ecosystem health. Nature 484:186–194CrossRefPubMedGoogle Scholar
  13. Gentleman R, Carey V, Bates D, Bolstad B, Dettling M, Dudoit S, Ellis B, Gautier L, Ge Y, Gentry J, Hornik K, Hothorn T, Huber W, Iacus S, Irizarry R, Leisch F, Li C, Maechler M, Rossini A, Sawitzki G, Smith C, Smyth G, Tierney L, Yang J, Zhang J (2004) Bioconductor: open software development for computational biology and bioinformatics. Genome Biol 5:R80CrossRefPubMedPubMedCentralGoogle Scholar
  14. Gijzen M, Nürnberger T (2006) Nep1-like proteins from plant pathogens: recruitment and diversification of the NPP1 domain across taxa. Phytochemistry 67:1800–1807CrossRefPubMedGoogle Scholar
  15. Giraldo MC, Dagdas YF, Gupta YK, Mentlak TA, Yi M, Martinez-Rocha AL, Saitoh H, Terauchi R, Talbot NJ, Valent B (2013) Two distinct secretion systems facilitate tissue invasion by the rice blast fungus Magnaporthe oryzae. Nat Commun 4:1996CrossRefPubMedPubMedCentralGoogle Scholar
  16. Godfrey D, Bohlenius H, Pedersen C, Zhang Z, Emmersen J, Thordal-Christensen H (2010) Powdery mildew and rust fungal effector candidates share N-terminal Y/F/WxC-motif. BMC Genom 11:317CrossRefGoogle Scholar
  17. Gowda M, Venu R, Raghupathy M, Nobuta K, Li H, Wing R, Stahlberg E, Couglan S, Haudenschild C, Dean R, Nahm B-H, Meyers B, Wang G-L (2006) Deep and comparative analysis of the mycelium and appressorium transcriptomes of Magnaporthe grisea using MPSS, RL-SAGE, and oligoarray methods. BMC Genom 7:310CrossRefGoogle Scholar
  18. Heath MC (2000) Nonhost resistance and nonspecific plant defenses. Curr Opin Plant Biol 3:315–319CrossRefPubMedGoogle Scholar
  19. Horton P, Park KJ, Obayashi T, Nakai K (2006) Protein subcellular localization prediction with WOLF PSORT. Ser Adv Bioinform 3:39–48Google Scholar
  20. Horton P, Park KJ, Obayashi T, Fujita N, Harada H, Adams-Collier CJ, Nakai K (2007) WoLF PSORT: protein localization predictor. Nucleic Acids Res 35:W585–W587CrossRefPubMedPubMedCentralGoogle Scholar
  21. Hyon G-S, Nga N, Chuma I, Inoue Y, Asano H, Murata N, Kusaba M, Tosa Y (2012) Characterization of interactions between barley and various host-specific subgroups of Magnaporthe oryzae and M. grisea. J Gen Plant Pathol 78:237–246CrossRefGoogle Scholar
  22. Jarosch B, Kogel KH, Schaffrath U (1999) The ambivalence of the barley Mlo locus: mutations conferring resistance against powdery mildew (Blumeria graminis f. sp. hordei) enhance susceptibility to the rice blast fungus Magnaporthe grisea. Mol Plant Microbe Interact 12:508–514CrossRefGoogle Scholar
  23. Jarosch B, Collins NC, Zellerhoff N, Schaffrath U (2005) RAR1, ROR1, and the actin cytoskeleton contribute to basal resistance to Magnaporthe grisea in barley. Mol Plant Microbe Interact 18:397–404CrossRefPubMedGoogle Scholar
  24. Jia Y, McAdams SA, Bryan GT, Hershey HP, Valent B (2000) Direct interaction of resistance gene and avirulence gene products confers rice blast resistance. EMBO J 19:4004–4014CrossRefPubMedPubMedCentralGoogle Scholar
  25. Jones JDG, Dangl JL (2006) The plant immune system. Nature 444:323–329CrossRefPubMedGoogle Scholar
  26. Kankanala P, Czymmek K, Valent B (2007) Roles for rice membrane dynamics and plasmodesmata during biotrophic invasion by the blast fungus. Plant Cell 19:706–724CrossRefPubMedPubMedCentralGoogle Scholar
  27. Kanzaki H, Yoshida K, Saitoh H, Fujisaki K, Hirabuchi A, Alaux L, Fournier E, Tharreau D, Terauchi R (2012) Arms race co-evolution of Magnaporthe oryzae AVR-Pik and rice Pik genes driven by their physical interactions. Plant J 72:894–907CrossRefPubMedGoogle Scholar
  28. Kershaw MJ, Talbot NJ (2009) Genome-wide functional analysis reveals that infection-associated fungal autophagy is necessary for rice blast disease. Proc Natl Acad Sci 106:15967–15972CrossRefPubMedPubMedCentralGoogle Scholar
  29. Khang CH, Berruyer R, Giraldo MC, Kankanala P, Park S-Y, Czymmek K, Kang S, Valent B (2010) Translocation of Magnaporthe oryzae effectors into rice cells and their subsequent cell-to-cell movement. Plant Cell 22:1388–1403CrossRefPubMedPubMedCentralGoogle Scholar
  30. Kleemann J, Rincon-Rivera LJ, Takahara H, Neumann U, van Themaat EVL, van der Does HC, Hacquard S, Stüber K, Will I, Schmalenbach W, Schmelzer E, O’Connell RJ (2012) Sequential delivery of host-induced virulence effectors by appressoria and intracellular hyphae of the phytopathogen Colletotrichum higginsianum. PLoS Pathog 8:e1002643CrossRefPubMedPubMedCentralGoogle Scholar
  31. Kulkarni RD, Kelkar HS, Dean RA (2003) An eight-cysteine-containing CFEM domain unique to a group of fungal membrane proteins. Trends Biochem Sci 28:118–121CrossRefPubMedGoogle Scholar
  32. Kulkarni R, Thon M, Pan H, Dean R (2005) Novel G-protein-coupled receptor-like proteins in the plant pathogenic fungus Magnaporthe grisea. Genome Biol 6:R24CrossRefPubMedPubMedCentralGoogle Scholar
  33. Kunoh H, Kuroda K, Hayashimoto A, Ishizaki H (1986) Induced susceptibility and enhanced resistance at the cellular level in barley coleoptiles. II. Timing and localization of induced susceptibility in a single coleoptile cell and its transfer to an adjacent cell. Can J Bot 64:889–895CrossRefGoogle Scholar
  34. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(T)(−Delta Delta C) method. Methods 25:402–408CrossRefPubMedGoogle Scholar
  35. Loehrer M, Botterweck J, Jahnke J, Mahlmann DM, Gaetgens J, Oldiges M, Horbach R, Deising H, Schaffrath U (2014) In vivo assessment by Mach-Zehnder double-beam interferometry of the invasive force exerted by the Asian soybean rust fungus (Phakopsora pachyrhizi). New Phytol 203:620–631CrossRefPubMedGoogle Scholar
  36. Lyngkjær MF, Carver TLW (1999) Induced accessibility and inaccessibility to Blumeria graminis f. sp hordei in barley epidermal cells attacked by a compatible isolate. Physiol Mol Plant Pathol 55:151–162CrossRefGoogle Scholar
  37. Lyngkjær MF, Carver TLW, Zeyen RJ (2001) Virulent Blumeria graminis infection induces penetration susceptibility and suppresses race-specific hypersensitive resistance against avirulent attack in Mla1-barley. Physiol Mol Plant Pathol 59:243–256CrossRefGoogle Scholar
  38. Mathioni S, Beló A, Rizzo C, Dean R, Donofrio N (2011) Transcriptome profiling of the rice blast fungus during invasive plant infection and in vitro stresses. BMC Genomics 12:1–20CrossRefGoogle Scholar
  39. Mathioni SM, Patel N, Riddick B, Sweigard JA, Czymmek KJ, Caplan JL, Kunjeti SG, Kunjeti S, Raman V, Hillman BI, Kobayashi DY, Donofrio NM (2013) Transcriptomics of the rice blast fungus Magnaporthe oryzae in response to the bacterial antagonist Lysobacter enzymogenes reveals candidate fungal defense response genes. PLoS One 8:e76487CrossRefPubMedPubMedCentralGoogle Scholar
  40. Mentlak TA, Kombrink A, Shinya T, Ryder LS, Otomo I, Saitoh H, Terauchi R, Nishizawa Y, Shibuya N, Thomma BPHJ, Talbot NJ (2012) Effector-mediated suppression of chitin-triggered immunity by Magnaporthe oryzae is necessary for rice blast disease. Plant Cell Online 24:322–335CrossRefGoogle Scholar
  41. Mosquera G, Giraldo MC, Khang CH, Coughlan S, Valent B (2009) Interaction transcriptome analysis identifies Magnaporthe oryzae BAS1-4 as biotrophy-associated secreted proteins in rice blast disease. Plant Cell 21:1273–1290CrossRefPubMedPubMedCentralGoogle Scholar
  42. Motteram J, Küfner I, Deller S, Brunner F, Hammond-Kosack KE, Nürnberger T, Rudd JJ (2009) Molecular characterization and functional analysis of MgNLP, the sole NPP1 domain-containing protein, from the fungal wheat leaf pathogen Mycosphaerella graminicola. Mol Plant Microbe Interact 22:790–799CrossRefPubMedGoogle Scholar
  43. Mysore KS, Ryu CM (2004) Nonhost resistance: How much do we know? Trends Plant Sci 9:97–104CrossRefPubMedGoogle Scholar
  44. Nga NTT, Hau VTB, Tosa Y (2009) Identification of genes for resistance to a Digitaria isolate of Magnaporthe grisea in common wheat cultivars. Genome 52:801–809CrossRefPubMedGoogle Scholar
  45. Nga NTT, Inoue Y, Chuma I, Hyon G-S, Okada K, Vy TTP, Kusaba M, Tosa Y (2012) Identification of a novel locus Rmo2 conditioning resistance in barley to host-specific subgroups of Magnaporthe oryzae. Phytopathology 102:674–682CrossRefPubMedGoogle Scholar
  46. Odenbach D, Breth B, Thines E, Weber RWS, Anke H, Foster AJ (2007) The transcription factor Con7p is a central regulator of infection-related morphogenesis in the rice blast fungus Magnaporthe grisea. Mol Microbiol 64:293–307CrossRefPubMedGoogle Scholar
  47. Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786CrossRefPubMedGoogle Scholar
  48. Pritchard L, Birch PRJ (2014) The zigzag model of plant–microbe interactions: is it time to move on? Mol Plant Pathol 15:865–870CrossRefPubMedGoogle Scholar
  49. Qutob D, Kemmerling B, Brunner F, Küfner I, Engelhardt S, Gust AA, Luberacki B, Seitz HU, Stahl D, Rauhut T, Glawischnig E, Schween G, Lacombe B, Watanabe N, Lam E, Schlichting R, Scheel D, Nau K, Dodt G, Hubert D, Gijzen M, Nürnberger T (2006) Phytotoxicity and innate immune responses induced by Nep1-like proteins. Plant Cell 18:3721–3744CrossRefPubMedPubMedCentralGoogle Scholar
  50. R Development Core Team (2009). http://www.R-project.org. Accessed 15 July 2009
  51. Rho HS, Kang S, Lee YH (2001) Agrobacterium tumefaciens-mediated transformation of the plant pathogenic fungus, Magnaporthe grisea. Mol Cells 12:407–411PubMedGoogle Scholar
  52. Saitoh H, Fujisawa S, Mitsuoka C, Ito A, Hirabuchi A, Ikeda K, Irieda H, Yoshino K, Yoshida K, Matsumura H, Tosa Y, Win J, Kamoun S, Takano Y, Terauchi R (2012) Large-scale gene disruption in Magnaporthe oryzae identifies MC69, a secreted protein required for infection by monocot and dicot fungal pathogens. PLoS Pathog 8:e1002711CrossRefPubMedPubMedCentralGoogle Scholar
  53. Schulze-Lefert P, Panstruga R (2011) A molecular evolutionary concept connecting nonhost resistance, pathogen host range, and pathogen speciation. Trends Plant Sci 16:117–125CrossRefPubMedGoogle Scholar
  54. Smyth GK (2004) Linear models and empirical bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol 3:1–25Google Scholar
  55. Sweigard JA, Carroll AM, Kang S, Farrall L, Chumley FG, Valent B (1995) Identification, cloning, and characterization of PWL2, a gene for host species specificity in the rice blast fungus. Plant Cell 7:1221–1233CrossRefPubMedPubMedCentralGoogle Scholar
  56. Talbot NJ (2003) On the trail of a cereal killer: investigating the biology of Magnaporthe grisea. Annu Rev Microbiol 57:177–202CrossRefPubMedGoogle Scholar
  57. Talbot NJ, Ebbole DJ, Hamer JE (1993) Identification and characterization of MPG1, a gene involved in pathogenicity from the rice blast fungus Magnaporthe grisea. Plant Cell 5:1575–1590CrossRefPubMedPubMedCentralGoogle Scholar
  58. Thordal-Christensen H (2003) Fresh insights into processes of nonhost resistance. Curr Opin Plant Biol 6:351–357CrossRefPubMedGoogle Scholar
  59. Tucker S, Orbach M (2007) Agrobacterium-mediated transformation to create an insertion library in Magnaporthe grisea. In: Ronald P (ed) Plant-pathogen interactions. Humana Press, New York, pp 57–68Google Scholar
  60. Untergasser A, Nijveen H, Rao X, Bisseling T, Geurts R, Leunissen JAM (2007) Primer3Plus, an enhanced web interface to Primer3. Nucleic Acids Res 35:W71–W74CrossRefPubMedPubMedCentralGoogle Scholar
  61. Voegele RT, Schmid A (2011) RT real-time PCR-based quantification of Uromyces fabae in planta. FEMS Microbiol Lett 322:131–137CrossRefPubMedGoogle Scholar
  62. Whisson SC, Boevink PC, Moleleki L, Avrova AO, Morales JG, Gilroy EM, Armstrong MR, Grouffaud S, van West P, Chapman S, Hein I, Toth IK, Pritchard L, Birch PRJ (2007) A translocation signal for delivery of oomycete effector proteins into host plant cells. Nature 450:115–118CrossRefPubMedGoogle Scholar
  63. Wilson RA, Talbot NJ (2009) Under pressure: investigating the biology of plant infection by Magnaporthe oryzae. Nat Rev Microbiol 7:185–195CrossRefPubMedGoogle Scholar
  64. Wu J, Kou Y, Bao J, Li Y, Tang M, Zhu X, Ponaya A, Xiao G, Li J, Li C, Song M-Y, Cumagun CJR, Deng Q, Lu G, Jeon JS, Naqvi NI, Zhou B (2015) Comparative genomics identifies the Magnaporthe oryzae avirulence effector AvrPi9 that triggers Pi9-mediated blast resistance in rice. New Phytologist 206:1463–1475CrossRefPubMedGoogle Scholar
  65. Yu J-H, Hamari Z, Han K-H, Seo J-A, Reyes-Domínguez Y, Scazzocchio C (2004) Double-joint PCR: a PCR-based molecular tool for gene manipulations in filamentous fungi. Fungal Genet Biol 41:973–981CrossRefPubMedGoogle Scholar
  66. Zellerhoff N, Jarosch B, Groenewald JZ, Crous PW, Schaffrath U (2006) Nonhost resistance of barley is successfully manifested against Magnaporthe grisea and a closely related Pennisetum-infecting lineage but is overcome by Magnaporthe oryzae. Mol Plant Microbe Interact 19:1014–1022CrossRefPubMedGoogle Scholar
  67. Zellerhoff N, Himmelbach A, Dong W, Bieri S, Schaffrath U, Schweizer P (2010) Nonhost resistance of barley to different fungal pathogens is associated with largely distinct, quantitative transcriptional responses. Plant Physiol 152:2053–2066CrossRefPubMedPubMedCentralGoogle Scholar
  68. Zhang S, Xu J-R (2014) Effectors and effector delivery in Magnaporthe oryzae. PLoS Pathog 10:e1003826CrossRefPubMedPubMedCentralGoogle Scholar
  69. Zhang H, Li D, Wang M, Liu J, Teng W, Cheng B, Huang Q, Wang M, Song W, Dong S, Zheng X, Zhang Z (2012) The Nicotiana benthamiana mitogen-activated protein kinase cascade and WRKY transcription factor participate in Nep1Mo-triggered plant responses. Mol Plant Microbe Interact 25:1639–1653CrossRefPubMedGoogle Scholar
  70. Zipfel C (2014) Plant pattern-recognition receptors. Trends Immunol 35:345–351CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Valerie Mogga
    • 1
  • Rhoda Delventhal
    • 1
  • Denise Weidenbach
    • 1
  • Samantha Langer
    • 1
  • Philipp M. Bertram
    • 1
  • Karsten Andresen
    • 2
  • Eckhard Thines
    • 2
    • 3
  • Thomas Kroj
    • 4
  • Ulrich Schaffrath
    • 1
  1. 1.Department of Plant PhysiologyRWTH Aachen UniversityAachenGermany
  2. 2.Institute of Biotechnology and Drug ResearchKaiserslauternGermany
  3. 3.BiotechnologyJohannes Gutenberg-UniversityMainzGermany
  4. 4.INRA, UMR BGPI, Campus International de BaillarguetMontpellier Cedex 5France

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