Increased Auxin Content and Altered Auxin Response in Barley Necrotic Mutant nec1

  • Anete Keisa
  • Ilva Nakurte
  • Laura Kunga
  • Liga Kale
  • Nils Rostoks
Conference paper


The role of hormone crosstalk in plant immunity is lately emerging as significant topic of plant physiology. Although crosstalk between salicylic acid and auxin affects plant disease resistance, molecular mechanisms of this process have not yet been uncovered in details. Mutations disrupting cyclic nucleotide-gated ion channel 4 (CNGC4) affect SA-mediated disease resistance in barley Hordeum vulgare and in A. thaliana. Significantly, decreased stomatal apertures of barley CNGC4 mutant nec1 and dwarfed stature of A. thaliana CNGC4 mutant dnd2 suggest that nonfunctional CNGC4 might be affecting also auxin signaling. Excised coleoptile elongation, stomatal conductance, and cell size measurements assaying physiological effect of exogenous auxin treatment suggested altered auxin signaling in nec1 mutant. Real-time qPCR analysis identified significant change in mRNA abundance of four auxin-related genes – YUCCA1, VT2, HVP1, and TIR1. Analysis of endogenous auxin content of nec1 plants detected ca. fourfold increase in indole acetic acid (IAA) content in nec1 leaves and roots compared to wt plants, as measured by HPLC. These results suggest that apart from SA-related disease resistance, CNGC4 functions also in auxin signaling in barley; therefore, barley nec1 mutant could serve as model system revealing role of SA-auxin crosstalk in plant disease resistance.


Barley nec1 mutant Lesion mimic mutant Auxin Disease resistance 



The study was funded by European Social Fund project 2009/0224/1DP/ and Latvian Council of Science grant Z-6142-090.


  1. Azevedo, C., Sadanandom, A., Kitagawa, K., Freialdenhoven, A., Shirasu, K., & Schulze-Lefert, P. (2002). The RAR1 interactor SGT1, an essential component of R gene-triggered disease resistance. Science, 295, 2073–2076.PubMedCrossRefGoogle Scholar
  2. Azevedo, C., Betsuyaku, S., Peart, J., Takahashi, A., Noël, L., Sadanandom, A., Casais, C., Parker, J., & Shirasu, K. (2006). Role of SGT1 in resistance protein accumulation in plant immunity. EMBO Journal, 25, 2007–2016.PubMedCrossRefGoogle Scholar
  3. Balague, C., Lin, B., Alcon, C., Flottes, G., Malmstrom, S., Kohler, C., Neuhaus, G., Pelletier, G., Gaymard, F., & Roby, D. (2003). HLM1, an essential signaling component in the hypersensitive response, is a member of the cyclic nucleotide-gated channel ion channel family. The Plant Cell, 15, 365–379.PubMedCrossRefGoogle Scholar
  4. Braun, N., Wyrzykowska, J., Muller, P., David, K., Couch, D., Perrot-Rechenmann, C., & Fleming, A. J. (2008). Conditional repression of AUXIN BINDING PROTEIN1 reveals that it coordinates cell division and cell expansion during postembryonic shoot development in Arabidopsis and tobacco. The Plant Cell, 20, 2746–2762.PubMedCrossRefGoogle Scholar
  5. Bright, J., Desikan, R., Hancock, J. T., Weir, I. S., & Neill, S. J. (2006). ABA-induced NO generation and stomatal closure in Arabidopsis are dependent on H2O2 synthesis. The Plant Journal, 45, 113–122.PubMedCrossRefGoogle Scholar
  6. Chen, Z., Agnew, J. L., Cohen, J. D., He, P., Shan, L., Sheen, J., & Kunkel, B. N. (2007). Pseudomonas syringae type III effector AvrRpt2 alters Arabidopsis thaliana auxin physiology. Proceedings of the National Academy of Sciences of the United States of America, 104, 20131–20136.PubMedCrossRefGoogle Scholar
  7. Cheng, N. H., Pittman, J. K., Barkla, B. J., Shigaki, T., & Hirschi, K. D. (2003). The Arabidopsis cax1 mutant exhibits impaired ion homeostasis, development, and hormonal responses and reveals interplay among vacuolar transporters. The Plant Cell, 15, 347–364.PubMedCrossRefGoogle Scholar
  8. Cheng, Y., Dai, X., & Zhao, Y. (2006). Auxin biosynthesis by the YUCCA flavin monooxygenases controls the formation of floral organs and vascular tissues in Arabidopsis. Genes & Development, 20, 1790–1799.CrossRefGoogle Scholar
  9. David, K. M., Couch, D., Braun, N., Brown, S., Grosclaude, J., & Perrot-Rechenmann, C. (2007). The auxin-binding protein 1 is essential for the control of cell cycle. The Plant Journal, 50, 197–206.PubMedCrossRefGoogle Scholar
  10. Dharmasiri, N., Dharmasiri, S., & Estelle, M. (2005). The F-box protein TIR1 is an auxin receptor. Nature, 435, 441–445.PubMedCrossRefGoogle Scholar
  11. Ding, X., Cao, Y., Huang, L., Zhao, J., Xu, C., Li, X., & Wang, S. (2008). Activation of the indole-3-acetic acid–amido synthetase GH3-8 suppresses expansin expression and promotes salicylate- and jasmonate-independent basal immunity in rice. The Plant Cell, 20, 228–240.PubMedCrossRefGoogle Scholar
  12. Dobrev, P. I., & Kaminek, M. (2002). Fast and efficient separation of cytokinins from auxin and abscisic acid and their purification using mixed-mode solid-phase extraction. Journal of Chromatography A, 950, 21–29.PubMedCrossRefGoogle Scholar
  13. Du, L., Ali, G. S., Simons, K. A., Hou, J., Yang, T., Reddy, A. S. N., & Poovaiah, B. W. (2009). Ca2+/calmodulin regulates salicylic-acid-mediated plant immunity. Nature, 457, 1154–1158.PubMedCrossRefGoogle Scholar
  14. Effendi, Y., Rietz, S., Fischer, U., & Scherer, G. F. E. (2011). The heterozygous abp1/ABP1 insertional mutant has defects in functions requiring polar auxin transport and in regulation of early auxin-regulated genes. The Plant Journal, 65, 282–294.PubMedCrossRefGoogle Scholar
  15. Fukuda, A., & Tanaka, Y. (2006). Effects of ABA, auxin, and gibberellin on the expression of genes for vacuolar H+ -inorganic pyrophosphatase, H+ -ATPase subunit A, and Na+/H+ antiporter in barley. Plant Physiology and Biochemistry, 44, 351–358.PubMedCrossRefGoogle Scholar
  16. Gaxiola, R. A., Palmgren, M. G., & Schumacher, K. (2007). Plant proton pumps. FEBS Letters, 581, 2204–2214.PubMedCrossRefGoogle Scholar
  17. Gehring, C. A., McConchie, R. M., Venis, M. A., & Parish, R. W. (1998). Auxin-binding-protein antibodies and peptides influence stomatal opening and alter cytoplasmic pH. Planta, 205, 581–586.PubMedCrossRefGoogle Scholar
  18. Glazebrook, J. (2005). Contrasting mechanisms of defense against biotrophic and necrotrophic pathogens. Annual Review of Phytopathology, 43, 205–227.PubMedCrossRefGoogle Scholar
  19. Grant, M. R., & Jones, J. D. G. (2009). Hormone (dis)harmony moulds plant health and disease. Science, 324, 750–752.PubMedCrossRefGoogle Scholar
  20. Gray, W. M., Muskett, P. R., Chuang, H., & Parker, J. E. (2003). Arabidopsis SGT1b is required for SCFTIR1-mediated auxin response. The Plant Cell, 15, 1310–1319.PubMedCrossRefGoogle Scholar
  21. Hesse, T., Feldwisch, J., Balshusemann, D., Bauw, G., Puype, M., Vandekerckhove, J., Lobler, M., Klambt, D., Schell, J., & Palme, K. (1989). Molecular cloning and structural analysis of a gene from Zea mays (L.) coding for a putative receptor for the plant hormone auxin. EMBO Journal, 8, 2453–2461.PubMedGoogle Scholar
  22. Iglesias, M. J., Terrile, M. C., & Casalongué, C. A. (2011). Auxin and salicylic acid signalings counteract during the adaptive response to stress. Plant Signaling & Behavior, 6, 452–454.CrossRefGoogle Scholar
  23. Jurkowski, G. I., Smith, R. K., Jr., Yu, I. C., Ham, J. H., Sharma, S. B., Klessig, D. F., Fengler, K. A., & Bent, A. F. (2004). Arabidopsis DND2, a second cyclic nucleotide-gated ion channel gene for which mutation causes the “defense, no death” phenotype. Molecular Plant-Microbe Interactions, 17, 511–520.PubMedCrossRefGoogle Scholar
  24. Kaplan, B., Sherman, T., & Fromm, H. (2007). Cyclic nucleotide-gated channels in plants. FEBS Letters, 581, 2237–2246.PubMedCrossRefGoogle Scholar
  25. Kazan, K., & Manners, J. M. (2009). Linking development to defense: auxin in plant–pathogen interactions. Trends in Plant Science, 14, 373–382.PubMedCrossRefGoogle Scholar
  26. Keisa, A., Kanberga-Silina, K., Nakurte, I., Kunga, L., & Rostoks, N. (2011). Differential disease resistance response in the barley necrotic mutant nec1. BMC Plant Biology, 11, 66.PubMedCrossRefGoogle Scholar
  27. Kepinski, S., & Leyser, O. (2005). The Arabidopsis F-box protein TIR1 is an auxin receptor. Nature, 435, 446–451.PubMedCrossRefGoogle Scholar
  28. Khazaie, H., Mohammady, S., Monneveux, P., & Stoddard, F. (2011). The determination of direct effect of carbon isotope discrimination (Δ), stomatal characteristics and water use efficiency on grain yield in wheat using sequential path analysis. Australian Journal of Crop Science, 5, 466–472.Google Scholar
  29. Kotake, T., Nakagawa, N., Takeda, K., & Sakurai, N. (2000). Auxin-induced elongation growth and expressions of cell wall-bound exo and endo-β-glucanases in barley coleoptiles. Plant & Cell Physiology, 41, 1272–1278.CrossRefGoogle Scholar
  30. Kriechbaumer, V., Park, W. J., Piotrowski, M., Meeley, R. B., Gierl, A., & Glawischnig, E. (2007). Maize nitrilases have a dual role in auxin homeostasis and b-cyanoalanine hydrolysis. Journal of Experimental Botany, 58, 4225–4233.PubMedCrossRefGoogle Scholar
  31. Kutschera, U. (2006). Acid growth and plant development. Science, 311, 952–954.PubMedCrossRefGoogle Scholar
  32. Livak, K. J., & Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCt method. Methods, 25, 402–408.PubMedCrossRefGoogle Scholar
  33. Lüthen, H., Bigdon, M., & Böttger, M. (1990). Reexamination of the acid growth theory of auxin action. Plant Physiology, 93, 931–939.PubMedCrossRefGoogle Scholar
  34. Ma, W., Qi, Z., Smigel, A., Walker, R. K., Verma, R., & Berkowitz, G. A. (2009). Ca2+, cAMP, and transduction of non-self perception during plant immune responses. Proceedings of the National Academy of Sciences of the United States of America, 106, 20995–21000.PubMedCrossRefGoogle Scholar
  35. Mashiguchi, K., Tanaka, K., Sakai, T., Sugawara, S., Kawaide, H., Natsume, M., Hanada, A., Yaeno, T., Shirasu, K., Yao, H., McSteen, P., Zhao, Y., Hayashi, K., Kamiya, Y., & Kasahar, H. (2011). The main auxin biosynthesis pathway in Arabidopsis. Proceedings of the National Academy of Sciences of the United States of America, 108, 18512–18517.PubMedCrossRefGoogle Scholar
  36. McSteen, P. (2010). Auxin and monocot development. Cold Spring Harbor Perspectives in Biology, 2, a001479.PubMedCrossRefGoogle Scholar
  37. Navarro, L., Dunoyer, P., Jay, F., Arnold, B., Dharmasiri, N., Estelle, M., Voinnet, O., & Jones, J. D. G. (2006). A plant miRNA contributes to antibacterial resistance by repressing auxin signaling. Science, 312, 436–439.PubMedCrossRefGoogle Scholar
  38. Perrot-Rechenmann, C. (2010). Cellular responses to auxin: Division versus expansion. Cold Spring Harbor Perspectives in Biology, 2, a001446.PubMedCrossRefGoogle Scholar
  39. Phillips, K. A., Skirpan, A. L., Liu, X., Christensen, A., Slewinski, T. L., HudsonC, Barazesh S., Cohen, J. D., Malcomber, S., & McSteen, P. (2011). vanishing tassel2 encodes a grass-specific tryptophan aminotransferase required for vegetative and reproductive development in maize. The Plant Cell, 23, 550–566.PubMedCrossRefGoogle Scholar
  40. Pieterse, C. M. J., Leon-Reyes, A., Van der Ent, S., & Van Wees, S. C. M. (2009). Networking by small-molecule hormones in plant immunity. Nature Chemical Biology, 5, 308–316.PubMedCrossRefGoogle Scholar
  41. Rostoks, N., Schmierer, D., Mudie, S., Drader, T., Brueggeman, R., Caldwell, D. G., Waugh, R., & Kleinhofs, A. (2006). Barley necrotic locus nec1 encodes the cyclic nucleotide-gated ion channel 4 homologous to the Arabidopsis HLM1. Molecular and General Genetics, 275, 159–168.Google Scholar
  42. Sakata, T., Oshino, T., Miura, S., Tomabechi, M., Tsunaga, Y., Higashitani, N., Miyazawa, Y., Takahashi, H., Watanabe, M., & Higashitani, A. (2010). Auxin reverse plant male sterility caused by high temperatures. Proceedings of the National Academy of Sciences of the United States of America, 107, 8569–8574.PubMedCrossRefGoogle Scholar
  43. Schenck, D., Christian, M., Jones, A., & Luthen, H. (2010). Rapid auxin-induced cell expansion and gene expression: a four-decade-old question revisited. Plant Physiology, 152, 1183–1185.PubMedCrossRefGoogle Scholar
  44. Spaepen, S., & Vanderleyden, J. (2011). Auxin and plant-microbe interactions. Cold Spring Harbor Perspectives in Biology, 3, a001438.PubMedCrossRefGoogle Scholar
  45. Spoel, S. H., & Dong, X. (2008). Making sense of hormone crosstalk during plant immune responses. Cell Host & Microbe, 3, 348–351.CrossRefGoogle Scholar
  46. Tan, X., Calderon-Villalobos, L. I., Sharon, M., Zheng, C., Robinson, C. V., Estelle, M., & Zheng, N. (2007). Mechanism of auxin perception by the TIR1 ubiquitin ligase. Nature, 446, 640–645.PubMedCrossRefGoogle Scholar
  47. Tanaka, Y., Sano, T., Tamaoki, M., Nakajima, N., Kondo, N., & Hasezawa, S. (2006). Cytokinin and auxin inhibit abscisic acid-induced stomatal closure by enhancing ethylene production in Arabidopsis. Journal of Experimental Botany, 57, 2259–2266.PubMedCrossRefGoogle Scholar
  48. Teale, W. D., Paponov, I. A., & Palme, K. (2006). Auxin in action: Signalling, transport and the control of plant growth and development. Nature Reviews Molecular Cell Biology, 7, 847–859.PubMedCrossRefGoogle Scholar
  49. Tromas, A., Braun, N., Muller, P., Khodus, T., Paponov, I. A., Palme, K., Ljung, K., Lee, J. Y., Benfey, P., Murray, J. A., Scheres, B., & Perrot-Rechenmann, C. (2009). The AUXIN BINDING PROTEIN 1 is required for differential auxin responses mediating root growth. PLoS One, 4, e6648.PubMedCrossRefGoogle Scholar
  50. Truman, W. M., Bennett, M. H., Turnbull, C. G. N., & Grant, M. R. (2010). Arabidopsis auxin mutants are compromised in systemic acquired resistance and exhibit aberrant accumulation of various indolic compounds. Plant Physiology, 152, 1562–1573.PubMedCrossRefGoogle Scholar
  51. Verhage, A., van Wees, S. C. M., & Pieterse, C. M. J. (2010). Plant immunity: It’s the hormones talking, but what do they say? Plant Physiology, 154, 536–540.PubMedCrossRefGoogle Scholar
  52. Wang, D., Pajerowska-Mukhtar, K., Hendrickson Culler, A., & Dong, X. (2007). Salicylic acid inhibits pathogen growth in plants through repression of the auxin signaling pathway. Current Biology, 17, 1784–1790.PubMedCrossRefGoogle Scholar
  53. Wang, Y., Gao, M., Li, Q., Wang, L., Wang, J., Jeon, J.-S., Qu, N., Zhang, Y., & He, Z. (2008). OsRAR1 and OsSGT1 physically interact and function in rice basal disease resistance. Molecular Plant-Microbe Interactions, 21, 294–303.PubMedCrossRefGoogle Scholar
  54. Won, C., Shen, X., Mashiguchi, K., Zheng, Z., Dai, X., Cheng, Y., Kasahara, H., Kamiya, Y., Choryc, J., & Zhao, Y. (2011). Conversion of tryptophan to indole-3-acetic acid by tryptophan aminotransferases of Arabidopsis and YUCCAs in Arabidopsis. Proceedings of the National Academy of Sciences of the United States of America, 108, 18518–18523.PubMedCrossRefGoogle Scholar
  55. Yang, T., & Poovaiah, B. W. (2000). Molecular and biochemical evidence for the involvement of calcium/calmodulin in auxin action. Journal of Biological Chemistry, 275, 3137–3143.PubMedCrossRefGoogle Scholar
  56. Zhang, X., Wang, H., Takemiya, A., Song, C., Kinoshita, T., & Shimazaki, K. (2004). Inhibition of blue light-dependent H+ pumping by abscisic acid through hydrogen peroxide-induced dephosphorylation of the plasma membrane H+-ATPase in guard cell protoplasts. Plant Physiology, 136, 4150–4158.PubMedCrossRefGoogle Scholar
  57. Zhang, J., Peng, Y., & Guo, Z. (2009a). Constitutive expression of pathogen-inducible OsWRKY31 enhances disease resistance and affects root growth and auxin response in transgenic rice plants. Cell Research, 18, 508–521.CrossRefGoogle Scholar
  58. Zhang, L., Lavery, L., Gill, U., Gill, K., Steffenson, B., Yan, G., Chen, X., & Kleinhofs, A. (2009b). A cation/proton-exchanging protein is a candidate for the barley NecS1 gene controlling necrosis and enhanced defense response to stem rust. Theoretical and Applied Genetics, 118, 385–397.PubMedCrossRefGoogle Scholar

Copyright information

© Zhejiang University Press and Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Anete Keisa
    • 1
  • Ilva Nakurte
    • 1
  • Laura Kunga
    • 1
  • Liga Kale
    • 1
  • Nils Rostoks
    • 1
  1. 1.Faculty of BiologyUniversity of LatviaRigaLatvia

Personalised recommendations