Bph6 encodes an exocyst-localized protein and confers broad resistance to planthoppers in rice

Article metrics

Abstract

The brown planthopper (BPH) and white-backed planthopper (WBPH) are the most destructive insect pests of rice, and they pose serious threats to rice production throughout Asia. Thus, there are urgent needs to identify resistance-conferring genes and to breed planthopper-resistant rice varieties. Here we report the map-based cloning and functional analysis of Bph6, a gene that confers resistance to planthoppers in rice. Bph6 encodes a previously uncharacterized protein that localizes to exocysts and interacts with the exocyst subunit OsEXO70E1. Bph6 expression increases exocytosis and participates in cell wall maintenance and reinforcement. A coordinated cytokinin, salicylic acid and jasmonic acid signaling pathway is activated in Bph6-carrying plants, which display broad resistance to all tested BPH biotypes and to WBPH without sacrificing yield, as these plants were found to maintain a high level of performance in a field that was heavily infested with BPH. Our results suggest that a superior resistance gene that evolved long ago in a region where planthoppers are found year round could be very valuable for controlling agricultural insect pests.

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: Map-based cloning of Bph6.
Fig. 2: Molecular characterization of Bph6.
Fig. 3: Demonstration that BPH6 interacts with exocyst subunit OsEXO70E1.
Fig. 4: Characterization of exocyst gene expression and cell walls in 9311-Bph6-NIL and 9311 plants.
Fig. 5: Analyses of phytohormones in Bph6-carrying plants.
Fig. 6: Characterization of Bph6-mediated resistance.
Fig. 7: Evolution of Bph6 alleles in rice.

References

  1. 1.

    Grist, D. H. & Lever, R. J. Pests of Rice (Longmans, Green and Co, London, 1969).

  2. 2.

    Cheng, X., Zhu, L. & He, G. Towards understanding of molecular interactions between rice and the brown planthopper. Mol. Plant 6, 621–634 (2013).

  3. 3.

    Sogawa, K. The rice brown planthopper: feeding physiology and host plant interactions. Annu. Rev. Entomol. 27, 49–73 (1982).

  4. 4.

    Sezer, M. & Butlin, R. K. The genetic basis of host plant adaptation in the brown planthopper (Nilaparvata lugens). Heredity 80, 499–508 (1998).

  5. 5.

    Dyck, V. A. & Thomas, B. in Brown Planthopper: Threat to Rice Production in Asia 3–20 (International Rice Research Institute, Manila, Philippines, 1979).

  6. 6.

    Catindig, J. L. A. et al. in Planthoppers: New Threats to the Sustainability of Intensive Rice Production Systems in Asia 191–220 (International Rice Research Institute, Manila, Philippines, 2009).

  7. 7.

    Matsumura, M. et al. in Planthoppers: New Threats to the Sustainability of Intensive Rice Production Systems in Asia 233–244 (International Rice Research Institute, Manila, Philippines, 2009).

  8. 8.

    Pathak, M. D., Cheng, C. H. & Fortuno, M. E. Resistance to Nephotettix impicticeps and Nilaparvata lugens in varieties of rice. Nature 223, 502–504 (1969).

  9. 9.

    Athwal, D. S., Pathak, M. D., Bacalangco, E. & Pura, C. D. Genetics of resistance to brown planthoppers and green leaf hoppers in Oryza sativa L. Crop Sci. 11, 747–750 (1971).

  10. 10.

    Ling, Y. & Weilin, Z. Genetic and biochemical mechanisms of rice resistance to planthopper. Plant Cell Rep. 35, 1559–1572 (2016).

  11. 11.

    Du, B. et al. Identification and characterization of Bph14, a gene conferring resistance to brown planthopper in rice. Proc. Natl. Acad. Sci. USA 106, 22163–22168 (2009).

  12. 12.

    Tamura, Y. et al. Map-based cloning and characterization of a brown planthopper resistance gene BPH26 from Oryza sativa L. ssp. indica cultivar ADR52. Sci. Rep. 4, 5872 (2014).

  13. 13.

    Cheng, X. et al. A rice lectin receptor-like kinase that is involved in innate immune responses also contributes to seed germination. Plant J. 76, 687–698 (2013).

  14. 14.

    Liu, Y. et al. A gene cluster encoding lectin receptor kinases confers broad-spectrum and durable insect resistance in rice. Nat. Biotechnol. 33, 301–305 (2015).

  15. 15.

    Wang, Y. et al. Map-based cloning and characterization of BPH29, a B3-domain-containing recessive gene conferring brown planthopper resistance in rice. J. Exp. Bot. 66, 6035–6045 (2015).

  16. 16.

    Zhao, Y. et al. Allelic diversity in an NLR gene BPH9 enables rice to combat planthopper variation. Proc. Natl. Acad. Sci. USA 113, 12850–12855 (2016).

  17. 17.

    Ren, J. et al. Bph32, a novel gene encoding an unknown SCR domain-containing protein, confers resistance against the brown planthopper in rice. Sci. Rep. 6, 37645 (2016).

  18. 18.

    Kabir, M. A. & Khush, G. S. Genetic analysis of resistance to brown planthopper resistance gene in rice. Euphytica 107, 23–28 (1988).

  19. 19.

    Qiu, Y., Guo, J., Jing, S., Zhu, L. & He, G. High-resolution mapping of the brown planthopper resistance gene Bph6 in rice and characterizing its resistance in the 9311 and Nipponbare near-isogenic backgrounds. Theor. Appl. Genet. 121, 1601–1611 (2010).

  20. 20.

    Wang, J. et al. EXPO, an exocyst-positive organelle distinct from multivesicular endosomes and autophagosomes, mediates cytosol-to-cell-wall exocytosis in Arabidopsis and tobacco cells. Plant Cell 22, 4009–4030 (2010).

  21. 21.

    Zárský, V., Kulich, I., Fendrych, M. & Pečenková, T. Exocyst complexes multiple functions in plant cells secretory pathways. Curr. Opin. Plant Biol. 16, 726–733 (2013).

  22. 22.

    Fujisaki, K. et al. Rice EXO70 interacts with a fungal effector, AVR-Pii, and is required for AVR-Pii-triggered immunity. Plant J. 83, 875–887 (2015).

  23. 23.

    Zhao, T. et al. A truncated NLR protein, TIR-NBS2, is required for activated defense responses in the exo70B1 mutant. PLoS Genet. 11, e1004945 (2015).

  24. 24.

    Kim, S. J. & Brandizzi, F. The plant secretory pathway: an essential factory for building the plant cell wall. Plant Cell Physiol. 55, 687–693 (2014).

  25. 25.

    Hao, P. et al. Herbivore-induced callose deposition on the sieve plates of rice: an important mechanism for host resistance. Plant Physiol. 146, 1810–1820 (2008).

  26. 26.

    Pieterse, C. M. J., Van der Does, D., Zamioudis, C., Leon-Reyes, A. & Van Wees, S. C. M. Hormonal modulation of plant immunity. Annu. Rev. Cell Dev. Biol. 28, 489–521 (2012).

  27. 27.

    McConn, M., Creelman, R. A., Bell, E., Mullet, J. E. & Browse, J. Jasmonate is essential for insect defense in Arabidopsis. Proc. Natl. Acad. Sci. USA 94, 5473–5477 (1997).

  28. 28.

    Van der Does, D. et al. Salicylic acid suppresses jasmonic acid signaling downstream of SCFCOI1-JAZ by targeting GCC promoter motifs via transcription factor ORA59. Plant Cell 25, 744–761 (2013).

  29. 29.

    Zhou, G. et al. Silencing OsHI-LOX makes rice more susceptible to chewing herbivores but enhances resistance to a phloem feeder. Plant J. 60, 638–648 (2009).

  30. 30.

    Guo, H. M., Li, H. C., Zhou, S. R., Xue, H. W. & Miao, X. X. Cis-12-oxo-phytodienoic acid stimulates rice defense response to a piercing–sucking insect. Mol. Plant 7, 1683–1692 (2014).

  31. 31.

    Howe, G. A. & Jander, G. Plant immunity to insect herbivores. Annu. Rev. Plant Biol. 59, 41–66 (2008).

  32. 32.

    Naseem, M., Wölfling, M. & Dandekar, T. Cytokinins for immunity beyond growth, galls and green islands. Trends Plant Sci. 19, 481–484 (2014).

  33. 33.

    Grosskinsky, D. K. et al. Cytokinins mediate resistance against Pseudomonas syringae in tobacco through increased antimicrobial phytoalexin synthesis independent of salicylic acid signaling. Plant Physiol. 157, 815–830 (2011).

  34. 34.

    Jiang, C. J. et al. Cytokinins act synergistically with salicylic acid to activate defense gene expression in rice. Mol. Plant Microbe Interact. 26, 287–296 (2013).

  35. 35.

    Argueso, C. T. et al. Two-component elements mediate interactions between cytokinin and salicylic acid in plant immunity. PLoS Genet. 8, e1002448 (2012).

  36. 36.

    Naseem, M., Kaltdorf, M. & Dandekar, T. The nexus between growth and defense signaling: auxin and cytokinin modulate plant immune response pathways. J. Exp. Bot. 66, 4885–4896 (2015).

  37. 37.

    Hart, S. V., Kogan, M. & Paxton, J. D. Effect of soybean phytoalexins on the herbivorous insects mexican bean beetle and soybean looper. J. Chem. Ecol. 9, 657–672 (1983).

  38. 38.

    Yamane, H. Biosynthesis of phytoalexins and regulatory mechanisms of it in rice. Biosci. Biotechnol. Biochem. 77, 1141–1148 (2013).

  39. 39.

    Ahuja, I., Kissen, R. & Bones, A. M. Phytoalexins in defense against pathogens. Trends Plant Sci. 17, 73–90 (2012).

  40. 40.

    Miyamoto, K. et al. Overexpression of the bZIP transcription factor OsbZIP79 suppresses the production of diterpenoid phytoalexin in rice cells. J. Plant Physiol. 173, 19–27 (2015).

  41. 41.

    Painter, R. H. in Insect Resistance in Crop Plants 23–83 (Macmillan, New York, 1951).

  42. 42.

    Sang, T. & Ge, S. Understanding rice domestication and implications for cultivar improvement. Curr. Opin. Plant Biol. 16, 139–146 (2013).

  43. 43.

    Fisher, M. C. et al. Emerging fungal threats to animal, plant and ecosystem health. Nature 484, 186–194 (2012).

  44. 44.

    Huang, X. et al. A map of rice genome variation reveals the origin of cultivated rice. Nature 490, 497–501 (2012).

  45. 45.

    Xu, X. et al. Resequencing 50 accessions of cultivated and wild rice yields markers for identifying agronomically important genes. Nat. Biotechnol. 30, 105–111 (2011).

  46. 46.

    Agrawal, A. A., Hastings, A. P., Johnson, M. T., Maron, J. L. & Salminen, J. P. Insect herbivores drive real-time ecological and evolutionary change in plant populations. Science 338, 113–116 (2012).

  47. 47.

    Yan, W. et al. Natural variation in Ghd7.1 plays an important role in grain yield and adaptation in rice. Cell Res. 23, 969–971 (2013).

  48. 48.

    Qiu, Y. F. et al. Identification of antibiosis and tolerance in rice varieties carrying brown planthopper resistance genes. Entomol. Exp. Appl. 141, 224–231 (2011).

  49. 49.

    Jing, S. et al. Development and use of EST-SSR markers for assessing genetic diversity in the brown planthopper (Nilaparvata lugens Stål). Bull. Entomol. Res. 102, 113–122 (2012).

  50. 50.

    Huang, Z., He, G. C., Shu, L. H., Li, X. H. & Zhang, Q. F. Identification and mapping of two brown planthopper resistance genes in rice. Theor. Appl. Genet. 102, 929–934 (2001).

  51. 51.

    Heinrichs, E., Medrano, F. & Rapusas, H. in Genetic Evaluation of Insect Resistance in Rice (International Rice Research Institute, Manila, Philippines, 1985).

  52. 52.

    Yang, Z., Chen, H., Tang, W., Hua, H. & Lin, Y. Development and characterzation of transgenic rice expressing two Bacillus thuringiensis genes. Pest Manag. Sci. 67, 414–422 (2011).

  53. 53.

    Walter, M. et al. Visualization of protein interactions in living plant cells using bimolecular fluorescence complementation. Plant J. 40, 428–438 (2004).

  54. 54.

    Chen, S., Songkumarn, P., Liu, J. & Wang, G. L. A versatile zero background T-vector system for gene cloning and functional genomics. Plant Physiol. 150, 1111–1121 (2009).

  55. 55.

    Zhang, Y. et al. A highly efficient rice green tissue protoplast system for transient gene expression and studying light/- and chloroplast-related processes. Plant Methods 7, 30 (2011).

  56. 56.

    Nelson, B. K., Cai, X. & Nebenführ, A. A multicolored set of in vivo organelle markers for colocalization studies in Arabidopsis and other plants. Plant J. 51, 1126–1136 (2007).

  57. 57.

    Weng, Q. M., Huang, Z., Wang, X. L., Zhu, L. L. & He, G. C. In situ localization of proteinase inhibitor mRNA in rice plant challenged with brown planthopper. Chin. Sci. Bull. 48, 979–982 (2003).

  58. 58.

    Pettolino, F. A., Walsh, C., Fincher, G. B. & Bacic, A. Determining the polysaccharide composition of plant cell walls. Nat. Protoc. 7, 1590–1607 (2012).

  59. 59.

    Dubois, M., Gilles, K. A., Hamilton, J. K., Robers, P. A. & Smith, F. Colorimetric method for determination of sugars and related substances. Anal. Chem. 28, 350–356 (1956).

  60. 60.

    Updegraff, D. M. Semimicro determination of cellulose in biological materials. Anal. Biochem. 32, 420–424 (1969).

  61. 61.

    Smyth, G. K. Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat. Appl. Genet. Mol. Biol. 3, e3 (2004).

  62. 62.

    Liu, C. et al. Revealing different systems responses to brown planthopper infestation for pest-susceptible and resistant rice plants with the combined metabonomic and gene expression analysis. J. Proteome Res. 9, 6774–6785 (2010).

  63. 63.

    Liu, H., Li, X., Xiao, J. & Wang, S. A convenient method for simultaneous quantification of multiple phytohormones and metabolites: application in study of rice–bacterium interaction. Plant Methods 8, 2 (2012).

  64. 64.

    Yuan, H. M., Liu, W. C. & Lu, Y. T. CATALASE2 coordinates SA-mediated repression of both auxin accumulation and JA biosynthesis in plant defenses. Cell Host Microbe 21, 143–155 (2017).

  65. 65.

    Librado, P. & Rozas, J. DnaSPv5: a software for comprehensive analysis of DNA polymorphism data. Bioinformatics 25, 1451–1452 (2009).

  66. 66.

    Tamura, K., Stecher, G., Peterson, D., Filipski, A. & Kumar, S. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol. Biol. Evol. 30, 2725–2729 (2013).

  67. 67.

    You, A. et al. Identification of quantitative trait loci across recombinant inbred lines and testcross populations for traits of agronomic importance in rice. Genetics 172, 1287–1300 (2006).

Download references

Acknowledgements

We thank Q. Qian (China National Rice Research Institute), L. Han (Chinese Academy of Agricultural Sciences), L. Yan (Jiangxi Academy of Agricultural Sciences) and D. Pan (Guangdong Academy of Agricultural Sciences) for kindly providing rice germplasm, S. Wang (Huazhong Agricultural University) for kindly providing the rice disease pathogen PXO145, Y. Lin (Huazhong Agricultural University) for kindly providing the striped stem borer insects, D. Zeng (China National Rice Research Institute) for kindly providing the WBPH insects, Y. Liu for suggestions for the experiments, and Q. Zhang, R. He and J. Blackwell for edits and suggestions. This work was supported by grants from the National Natural Science Foundation of China (31230060 and 31630063, both to G.H.), the National Program on Research and Development of Transgenic Plants (2016ZX08009-003-001 to G.H.) and the National Key Research and Development Program (2016YFD0100600 and 2016YFD0100900, both to G.H.).

Author information

G.H. conceived and supervised the project; G.H. and J.G. designed the experiments; J.G. performed most of the experiments; C.X., Y.Z., D.W., B.D., X.W., Y.O., X.L., W.W., Y.Q., S.J., B.C., X.S., H.W., Y.M., Y.W., L.H., S.S., L.Z., X.X., R.C. and Y.F. performed some of the experiments; and J.G., C.X., D.W., B.D., Y.Z. and G.H. analyzed data and wrote the manuscript.

Correspondence to Bo Du or Guangcun He.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Additional information

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

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–27 and Supplementary Tables 1, 2, 4, 5, 7, 10 and 11

Life Sciences Reporting Summary

Supplementary Table 3

GeneChip data for comparing Nip-Bph6-NIL and Bph6-RNAi plants

Supplementary Table 6

Information on accessions of wild rice and cultivated varieties that were sequenced in the Bph6 coding region

Supplementary Table 8

List of SNPs between Swarnalata or Nipponbare and the 80 haplotypes

Supplementary Table 9

List of indels between Swarnalata or Nipponbare and the 80 haplotypes

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Further reading