Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Polymorphic residues in rice NLRs expand binding and response to effectors of the blast pathogen

A Publisher Correction to this article was published on 20 August 2018

This article has been updated

Abstract

Accelerated adaptive evolution is a hallmark of plant–pathogen interactions. Plant intracellular immune receptors (NLRs) often occur as allelic series with differential pathogen specificities. The determinants of this specificity remain largely unknown. Here, we unravelled the biophysical and structural basis of expanded specificity in the allelic rice NLR Pik, which responds to the effector AVR-Pik from the rice blast pathogen Magnaporthe oryzae. Rice plants expressing the Pikm allele resist infection by blast strains expressing any of three AVR-Pik effector variants, whereas those expressing Pikp only respond to one. Unlike Pikp, the integrated heavy metal-associated (HMA) domain of Pikm binds with high affinity to each of the three recognized effector variants, and variation at binding interfaces between effectors and Pikp-HMA or Pikm-HMA domains encodes specificity. By understanding how co-evolution has shaped the response profile of an allelic NLR, we highlight how natural selection drove the emergence of new receptor specificities. This work has implications for the engineering of NLRs with improved utility in agriculture.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: The Pikm-mediated cell death response to AVR-Pik effector variants in N.benthamiana phenocopies the Pikm resistance profile in rice.
Fig. 2: Different affinities underpin the recognition and response of Pik NLR alleles to AVR-Pik effector variants.
Fig. 3: Structures of Pikm-HMA in complex with AVR-Pik effectors.
Fig. 4: Different interactions at interface 3 in the complexes of Pikm-HMA and Pikp-HMA with AVR-PikD support recognition and response.
Fig. 5: Altered interactions across the interfaces of Pikp-HMA with AVR-PikD and AVR-PikE underpin the differences in recognition and response.
Fig. 6: Mutations at different interfaces in the Pik-HMA–effector complexes have differential effects on interactions and phenotypes.

Change history

  • 20 August 2018

    In the version of this Article originally published, in Fig. 1b the single-letter code for the amino acid polymorphism at position 46 in the schematic of the AVR-PikE variant was incorrectly given as ‘H’. The correct amino acid is ‘N’. This has now been amended in all versions of the Article.

References

  1. 1.

    Jones, J. D., Vance, R. E. & Dangl, J. L. Intracellular innate immune surveillance devices in plants and animals. Science 354, aaf6395 (2016).

    Article  PubMed  CAS  Google Scholar 

  2. 2.

    Ronald, P. C. & Beutler, B. Plant and animal sensors of conserved microbial signatures. Science 330, 1061–1064 (2010).

    Article  PubMed  CAS  Google Scholar 

  3. 3.

    Dodds, P. N. & Rathjen, J. P. Plant immunity: towards an integrated view of plant–pathogen interactions. Nat. Rev. Genet. 11, 539–548 (2010).

    Article  PubMed  CAS  Google Scholar 

  4. 4.

    Win, J. et al. Effector biology of plant-associated organisms: concepts and perspectives. Cold Spring Harb. Symp. Quant. Biol. 77, 235–247 (2012).

    Article  PubMed  CAS  Google Scholar 

  5. 5.

    Bialas, A. et al. Lessons in effector and NLR biology of plant–microbe systems. Mol. Plant Microbe Interact. 31, 34–45 (2017).

    Article  PubMed  Google Scholar 

  6. 6.

    Ellis, J. G., Lawrence, G. J., Luck, J. E. & Dodds, P. N. Identification of regions in alleles of the flax rust resistance gene L that determine differences in gene-for-gene specificity. Plant Cell 11, 495–506 (1999).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  7. 7.

    Allen, R. L. et al. Host–parasite coevolutionary conflict between Arabidopsis and downy mildew. Science 306, 1957–1960 (2004).

    Article  PubMed  CAS  Google Scholar 

  8. 8.

    Bhullar, N. K., Zhang, Z., Wicker, T. & Keller, B. Wheat gene bank accessions as a source of new alleles of the powdery mildew resistance gene Pm3: a large scale allele mining project. BMC Plant Biol. 10, 88 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  9. 9.

    Seeholzer, S. et al. Diversity at the Mla powdery mildew resistance locus from cultivated barley reveals sites of positive selection. Mol. Plant Microbe Interact. 23, 497–509 (2010).

    Article  PubMed  CAS  Google Scholar 

  10. 10.

    Srichumpa, P., Brunner, S., Keller, B. & Yahiaoui, N. Allelic series of four powdery mildew resistance genes at the Pm3 locus in hexaploid bread wheat. Plant Physiol. 139, 885–895 (2005).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  11. 11.

    Lu, X. et al. Allelic barley MLA immune receptors recognize sequence-unrelated avirulence effectors of the powdery mildew pathogen. Proc. Natl Acad. Sci. USA 113, E6486–E6495 (2016).

    Article  PubMed  CAS  Google Scholar 

  12. 12.

    Dodds, P. N. et al. Direct protein interaction underlies gene-for-gene specificity and coevolution of the flax resistance genes and flax rust avirulence genes. Proc. Natl Acad. Sci. USA 103, 8888–8893 (2006).

    Article  PubMed  CAS  Google Scholar 

  13. 13.

    Krasileva, K. V., Dahlbeck, D. & Staskawicz, B. J. Activation of an Arabidopsis resistance protein is specified by the in planta association of its leucine-rich repeat domain with the cognate oomycete effector. Plant Cell 22, 2444–2458 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  14. 14.

    Steinbrenner, A. D., Goritschnig, S. & Staskawicz, B. J. Recognition and activation domains contribute to allele-specific responses of an Arabidopsis NLR receptor to an oomycete effector protein. PLoS Pathog. 11, e1004665 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. 15.

    Wang, C. I. et al. Crystal structures of flax rust avirulence proteins AvrL567-A and -D reveal details of the structural basis for flax disease resistance specificity. Plant Cell 19, 2898–2912 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  16. 16.

    Huang, J., Si, W., Deng, Q., Li, P. & Yang, S. Rapid evolution of avirulence genes in rice blast fungus Magnaporthe oryzae. BMC Genet. 15, 45 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  17. 17.

    Raffaele, S. et al. Genome evolution following host jumps in the Irish potato famine pathogen lineage. Science 330, 1540–1543 (2010).

    Article  PubMed  CAS  Google Scholar 

  18. 18.

    Yoshida, K. et al. Association genetics reveals three novel avirulence genes from the rice blast fungal pathogen Magnaporthe oryzae. Plant Cell 21, 1573–1591 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  19. 19.

    Dangl, J. L., Horvath, D. M. & Staskawicz, B. J. Pivoting the plant immune system from dissection to deployment. Science 341, 746–751 (2013).

    Article  PubMed  CAS  Google Scholar 

  20. 20.

    Rodriguez-Moreno, L., Song, Y. & Thomma, B. P. Transfer and engineering of immune receptors to improve recognition capacities in crops. Curr. Opin. Plant Biol. 38, 42–49 (2017).

    Article  PubMed  CAS  Google Scholar 

  21. 21.

    Eitas, T. K. & Dangl, J. L. NB-LRR proteins: pairs, pieces, perception, partners, and pathways. Curr. Opin. Plant Biol. 13, 472–477 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  22. 22.

    Wu, C. H., Belhaj, K., Bozkurt, T. O., Birk, M. S. & Kamoun, S. Helper NLR proteins NRC2a/b and NRC3 but not NRC1 are required for Pto-mediated cell death and resistance in Nicotiana benthamiana. New Phytol. 209, 1344–1352 (2016).

    Article  PubMed  Google Scholar 

  23. 23.

    Narusaka, M. et al. RRS1 and RPS4 provide a dual Resistance-gene system against fungal and bacterial pathogens. Plant J. 60, 218–226 (2009).

    Article  PubMed  CAS  Google Scholar 

  24. 24.

    Sinapidou, E. et al. Two TIR:NB:LRR genes are required to specify resistance to Peronospora parasitica isolate Cala2 in Arabidopsis. Plant J. 38, 898–909 (2004).

    Article  PubMed  CAS  Google Scholar 

  25. 25.

    Ashikawa, I. et al. Two adjacent nucleotide-binding site-leucine-rich repeat class genes are required to confer Pikm-specific rice blast resistance. Genetics 180, 2267–2276 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  26. 26.

    Lee, S. K. et al. Rice Pi5-mediated resistance to Magnaporthe oryzae requires the presence of two coiled-coil-nucleotide-binding-leucine-rich repeat genes. Genetics 181, 1627–1638 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  27. 27.

    Wu, C. H. et al. NLR network mediates immunity to diverse plant pathogens. Proc. Natl Acad. Sci. USA 114, 8113–8118 (2017).

    PubMed  CAS  Google Scholar 

  28. 28.

    Cesari, S., Bernoux, M., Moncuquet, P., Kroj, T. & Dodds, P. N. A novel conserved mechanism for plant NLR protein pairs: the “integrated decoy” hypothesis. Front. Plant Sci. 5, 606 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Kroj, T., Chanclud, E., Michel-Romiti, C., Grand, X. & Morel, J. B. Integration of decoy domains derived from protein targets of pathogen effectors into plant immune receptors is widespread. New Phytol. 210, 618–626 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  30. 30.

    Sarris, P. F., Cevik, V., Dagdas, G., Jones, J. D. & Krasileva, K. V. Comparative analysis of plant immune receptor architectures uncovers host proteins likely targeted by pathogens. BMC Biol. 14, 8 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  31. 31.

    Wu, C. H., Krasileva, K. V., Banfield, M. J., Terauchi, R. & Kamoun, S.The “sensor domains” of plant NLR proteins: more than decoys?. Front. Plant Sci. 6, 134 (2015).

    PubMed  PubMed Central  Google Scholar 

  32. 32.

    Okuyama, Y. et al. A multifaceted genomics approach allows the isolation of the rice Pia-blast resistance gene consisting of two adjacent NBS-LRR protein genes. Plant J. 66, 467–479 (2011).

    Article  PubMed  CAS  Google Scholar 

  33. 33.

    Maqbool, A. et al. Structural basis of pathogen recognition by an integrated HMA domain in a plant NLR immune receptor. eLife 4, e08709 (2015).

    Article  PubMed Central  Google Scholar 

  34. 34.

    Kanzaki, H. et al. Arms race co-evolution of Magnaporthe oryzae AVR-Pik and rice Pik genes driven by their physical interactions. Plant J. 72, 894–907 (2012).

    Article  PubMed  CAS  Google Scholar 

  35. 35.

    Costanzo, S. & Jia, Y. L. Sequence variation at the rice blast resistance gene Pi-km locus: implications for the development of allele specific markers. Plant Sci. 178, 523–530 (2010).

    Article  CAS  Google Scholar 

  36. 36.

    Krissinel, E. Stock-based detection of protein oligomeric states in jsPISA. Nucleic Acids Res. 43, W314–W319 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  37. 37.

    de Guillen, K. et al. Structure analysis uncovers a highly diverse but structurally conserved effector family in phytopathogenic fungi. PLoS Pathog. 11, e1005228 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. 38.

    Bent, A. F. et al. RPS2 of Arabidopsis thaliana: a leucine-rich repeat class of plant disease resistance genes. Science 265, 1856–1860 (1994).

    Article  PubMed  CAS  Google Scholar 

  39. 39.

    Mindrinos, M., Katagiri, F., Yu, G. L. & Ausubel, F. M. The A. thaliana disease resistance gene RPS2 encodes a protein containing a nucleotide-binding site and leucine-rich repeats. Cell 78, 1089–1099 (1994).

    Article  PubMed  CAS  Google Scholar 

  40. 40.

    Whitham, S. et al. The product of the tobacco mosaic virus resistance gene N: similarity to toll and the interleukin-1 receptor. Cell 78, 1101–1115 (1994).

    Article  PubMed  CAS  Google Scholar 

  41. 41.

    Le Roux, C. et al. A receptor pair with an integrated decoy converts pathogen disabling of transcription factors to immunity. Cell 161, 1074–1088 (2015).

    Article  PubMed  CAS  Google Scholar 

  42. 42.

    Sarris, P. F. et al. A plant immune receptor detects pathogen effectors that target WRKY transcription factors. Cell 161, 1089–1100 (2015).

    Article  PubMed  CAS  Google Scholar 

  43. 43.

    Zhang, Z. M. et al. Mechanism of host substrate acetylation by a YopJ family effector. Nat. Plants 3, 17115 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  44. 44.

    Ortiz, D. et al. Recognition of the Magnaporthe oryzae effector AVR-Pia by the decoy domain of the rice NLR immune receptor RGA5. Plant Cell 29, 156–168 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  45. 45.

    Lobstein, J. et al. SHuffle, a novel Escherichia coli protein expression strain capable of correctly folding disulfide bonded proteins in its cytoplasm. Microb. Cell Fact. 11, 56 (2012).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  46. 46.

    Studier, F. W. Protein production by auto-induction in high density shaking cultures. Protein Expr. Purif. 41, 207–234 (2005).

    Article  PubMed  CAS  Google Scholar 

  47. 47.

    Wickham, H. ggplot. Elegant Graphics for Data Analysis (Springer, New York, NY, 2009).

  48. 48.

    Winter, G. xia2: An expert system for macromolecular crystallography data reduction. J. Appl. Crystallogr. 43, 186–190 (2010).

    Article  CAS  Google Scholar 

  49. 49.

    Evans, P. R. & Murshudov, G. N. How good are my data and what is the resolution? Acta Crystallogr. D Biol. Crystallogr. 69, 1204–1214 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  50. 50.

    Winn, M. D. et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. D Biol. Crystallogr. 67, 235–242 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  51. 51.

    McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  52. 52.

    Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  53. 53.

    Murshudov, G. N. et al. REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr. D Biol. Crystallogr. 67, 355–367 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  54. 54.

    Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D Biol. Crystallogr. 66, 12–21 (2010).

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by the BBSRC (grants BB/J00453, BB/P012574 and BB/M02198X), the ERC (proposal 743165), the John Innes Foundation, the Gatsby Charitable Foundation and JSPS KAKENHI 15H05779. We thank the Diamond Light Source (beamlines I03 and I04 under proposals MX9475 and MX13467) for access to X-ray data collection facilities. We also thank D. Lawson and C. Stevenson (JIC X-ray Crystallography/Biophysical Analysis Platform) for help with protein structure determination and SPR.

Author information

Affiliations

Authors

Contributions

J.C.D.l.C. and M.F. performed all of the experiments. J.C.D.l.C., M.F. and M.J.B. designed the experiments and analysed the data. A.M. and H.S. assisted with construct design and the initial protein production. R.T. and S.K. analysed the data. J.C.D.l.C., M.F. and M.J.B. wrote the manuscript with input from all authors.

Corresponding author

Correspondence to Mark J. Banfield.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary Methods, Supplementary Figures 1–5, Supplementary Tables 1–4 and Supplementary References

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

De la Concepcion, J.C., Franceschetti, M., Maqbool, A. et al. Polymorphic residues in rice NLRs expand binding and response to effectors of the blast pathogen. Nature Plants 4, 576–585 (2018). https://doi.org/10.1038/s41477-018-0194-x

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing