, Volume 242, Issue 6, pp 1361–1390 | Cite as

Quantitative proteomics analysis reveals that S-nitrosoglutathione reductase (GSNOR) and nitric oxide signaling enhance poplar defense against chilling stress

  • Tielong Cheng
  • Jinhui Chen
  • Abd_Allah EF
  • Pengkai Wang
  • Guangping Wang
  • Xiangyang Hu
  • Jisen ShiEmail author
Original Article


Main conclusion

NO acts as the essential signal to enhance poplar tolerance to chilling stress via antioxidant enzyme activities and protein S -nitrosylation modification, NO signal is also strictly controlled by S -nitrosoglutathione reductase and nitrate reductase to avoid the over-accumulation of reactive nitrogen species.

Poplar (Populus trichocarpa) are fast growing woody plants with both ecological and economic value; however, the mechanisms by which poplar adapts to environmental stress are poorly understood. In this study, we used isobaric tags for relative and absolute quantification proteomic approach to characterize the response of poplar exposed to cold stress. We identified 114 proteins that were differentially expressed in plants exposed to cold stress. In particular, some of the proteins are involved in reactive oxygen species (ROS) and reactive nitrogen species (RNS) metabolism. Further physiological analysis showed that nitric oxide (NO) signaling activated a series of downstream defense responses. We further demonstrated that NO activated antioxidant enzyme activities and S-nitrosoglutathione reductase (GSNOR) activities, which would reduce ROS and RNS toxicity and thereby enhance poplar tolerance to cold stress. Suppressing NO accumulation or GSNOR activity aggravated cold damage to poplar leaves. Moreover, our results showed that RNS can suppress the activities of GSNOR and NO nitrate reductase (NR) by S-nitrosylation to fine-tune the NO signal and modulate ROS levels by modulating the S-nitrosylation of ascorbate peroxidase protein. Hence, our data demonstrate that NO signaling activates multiple pathways that enhance poplar tolerances to cold stress, and that NO signaling is strictly controlled through protein post-translational modification by S-nitrosylation.


Cold stress Nitric oxide Poplar Proteome S-Nitrosoglutathione reductase 



Abscisic acid


Ascorbate peroxidase


2-(4-Carboxyphenyl)-4, 4, 5, 5–tetramethylimidazoline-1-oxyl-3-oxide


Dodecanoic acid


Dehydroascorbate reductase


Glutathione reductase




GSNO reductase




Nitric oxide


NO synthase


Nitrate reductase


Jasmonic acid


Monodehydroascorbate reductase


Reactive nitrogen species


Reactive oxygen species





The authors would like to extend their sincere appreciation to the Deanship of Scientific Research at King Saud University for funding this research (Research Group NO. RG 1435-014).

Supplementary material

425_2015_2374_MOESM1_ESM.xls (204 kb)
Suppl. Table S1 The detail information on the identified protein (XLS 204 kb)


  1. Able AJ (2003) Role of reactive oxygen species in the response of barley to necrotrophic pathogens. Protoplasma 221:137–143CrossRefPubMedGoogle Scholar
  2. Alexandersson E, Ali A, Resjo S, Andreasson E (2013) Plant secretome proteomics. Front. Plant Sci 4:9Google Scholar
  3. An F, Zhang X, Zhu Z, Ji Y, He W, Jiang Z, Li M, Guo H (2012) Coordinated regulation of apical hook development by gibberellins and ethylene in etiolated Arabidopsis seedlings. Cell Res 22:915–927CrossRefPubMedPubMedCentralGoogle Scholar
  4. 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 72:248–254CrossRefPubMedGoogle Scholar
  5. Bai X, Yang L, Tian M, Chen J, Shi J, Yang Y, Hu X (2011a) Nitric oxide enhances desiccation tolerance of recalcitrant Antiaris toxicaria seeds via protein S-nitrosylation and carbonylation. PLoS One 6:e20714CrossRefPubMedPubMedCentralGoogle Scholar
  6. Bai X, Yang L, Yang Y, Ahmad P, Yang Y, Hu X (2011b) Deciphering the protective role of nitric oxide against salt stress at the physiological and proteomic levels in maize. J Proteome Res 10:4349–4364CrossRefPubMedGoogle Scholar
  7. Bai XG, Chen JH, Kong XX, Todd CD, Yang YP, Hu XY, Li DZ (2012) Carbon monoxide enhances the chilling tolerance of recalcitrant Baccaurea ramiflora seeds via nitric oxide-mediated glutathione homeostasis. Free Radic Biol Med 53:710–720CrossRefPubMedGoogle Scholar
  8. Baron KN, Schroeder DF, Stasolla C (2012) Transcriptional response of abscisic acid (ABA) metabolism and transport to cold and heat stress applied at the reproductive stage of development in Arabidopsis thaliana. Plant Sci 188–189:48–59CrossRefPubMedGoogle Scholar
  9. Barroso JB, Corpas FJ, Carreras A, Rodriguez-Serrano M, Esteban FJ, Fernandez-Ocana A, Chaki M, Romero-Puertas MC, Valderrama R, Sandalio LM, del Rio LA (2006) Localization of S-nitrosoglutathione and expression of S-nitrosoglutathione reductase in pea plants under cadmium stress. J Exp Bot 57:1785–1793CrossRefPubMedGoogle Scholar
  10. Beck EH, Heim R, Hansen J (2004) Plant resistance to cold stress: mechanisms and environmental signals triggering frost hardening and dehardening. J Biosci 29:449–459CrossRefPubMedGoogle Scholar
  11. Bindschedler LV, Cramer R (2011) Quantitative plant proteomics. Proteomics 11:756–775CrossRefPubMedGoogle Scholar
  12. Catala R, Lopez-Cobollo R, Mar Castellano M, Angosto T, Alonso JM, Ecker JR, Salinas J (2014) The Arabidopsis 14-3-3 protein RARE COLD INDUCIBLE 1A links low-temperature response and ethylene biosynthesis to regulate freezing tolerance and cold acclimation. Plant Cell 26:3326–3342CrossRefPubMedPubMedCentralGoogle Scholar
  13. Chaki M, Fernandez-Ocana AM, Valderrama R, Carreras A, Esteban FJ, Luque F, Gomez-Rodriguez MV, Begara-Morales JC, Corpas FJ, Barroso JB (2009) Involvement of reactive nitrogen and oxygen species (RNS and ROS) in sunflower-mildew interaction. Plant Cell Physiol 50:265–279CrossRefPubMedGoogle Scholar
  14. Chaki M, Valderrama R, Fernandez-Ocana AM, Carreras A, Gomez-Rodriguez MV, Pedrajas JR, Begara-Morales JC, Sanchez-Calvo B, Luque F, Leterrier M, Corpas FJ, Barroso JB (2011) Mechanical wounding induces a nitrosative stress by down-regulation of GSNO reductase and an increase in S-nitrosothiols in sunflower (Helianthus annuus) seedlings. J Exp Bot 62:1803–1813CrossRefPubMedPubMedCentralGoogle Scholar
  15. Chen LT, Wu K (2010) Role of histone deacetylases HDA6 and HDA19 in ABA and abiotic stress response. Plant Signal Behav 5:1318–1320CrossRefPubMedPubMedCentralGoogle Scholar
  16. Chen Q, Yang L, Ahmad P, Wan X, Hu X (2011) Proteomic profiling and redox status alteration of recalcitrant tea (Camellia sinensis) seed in response to desiccation. Planta 233:583–592CrossRefPubMedGoogle Scholar
  17. Chinnusamy V, Gong Z, Zhu JK (2008) Abscisic acid-mediated epigenetic processes in plant development and stress responses. J Integr Plant Biol 50:1187–1195CrossRefPubMedPubMedCentralGoogle Scholar
  18. Corpas FJ, Chaki M, Fernandez-Ocana A, Valderrama R, Palma JM, Carreras A, Begara-Morales JC, Airaki M, del Rio LA, Barroso JB (2008) Metabolism of reactive nitrogen species in pea plants under abiotic stress conditions. Plant Cell Physiol 49:1711–1722CrossRefPubMedGoogle Scholar
  19. Desikan R, Cheung MK, Bright J, Henson D, Hancock JT, Neill SJ (2004) ABA, hydrogen peroxide and nitric oxide signalling in stomatal guard cells. J Exp Bot 55:205–212CrossRefPubMedGoogle Scholar
  20. Ding Y, Li H, Zhang X, Xie Q, Gong Z, Yang S (2015) OST1 kinase modulates freezing tolerance by enhancing ICE1 stability in Arabidopsis. Dev Cell 32:278–289CrossRefPubMedGoogle Scholar
  21. Dong CH, Agarwal M, Zhang Y, Xie Q, Zhu JK (2006) The negative regulator of plant cold responses, HOS1, is a RING E3 ligase that mediates the ubiquitination and degradation of ICE1. Proc Natl Acad Sci USA 103:8281–8286CrossRefPubMedPubMedCentralGoogle Scholar
  22. Feechan A, Kwon E, Yun BW, Wang Y, Pallas JA, Loake GJ (2005) A central role for S-nitrosothiols in plant disease resistance. Proc Natl Acad Sci USA 102:8054–8059CrossRefPubMedPubMedCentralGoogle Scholar
  23. Forrester MT, Foster MW, Benhar M, Stamler JS (2009) Detection of protein S-nitrosylation with the biotin-switch technique. Free Radic Biol Med 46:119–126CrossRefPubMedPubMedCentralGoogle Scholar
  24. Foyer CH, Noctor G (2011) Ascorbate and glutathione: the heart of the redox hub. Plant Physiol 155:2–18CrossRefPubMedPubMedCentralGoogle Scholar
  25. Fragoso V, Rothe E, Baldwin IT, Kim SG (2014) Root jasmonic acid synthesis and perception regulate folivore-induced shoot metabolites and increase Nicotiana attenuata resistance. New Phytol 202:1335–1345CrossRefPubMedGoogle Scholar
  26. Frungillo L, Skelly MJ, Loake GJ, Spoel SH, Salgado I (2014) S-nitrosothiols regulate nitric oxide production and storage in plants through the nitrogen assimilation pathway. Nat Commun 5:5401CrossRefPubMedPubMedCentralGoogle Scholar
  27. Gaudet M, Pietrini F, Beritognolo I, Iori V, Zacchini M, Massacci A, Mugnozza GS, Sabatti M (2011) Intraspecific variation of physiological and molecular response to cadmium stress in Populus nigra L. Tree Physiol 31:1309–1318CrossRefPubMedGoogle Scholar
  28. He Y, Tang RH, Hao Y, Stevens RD, Cook CW, Ahn SM, Jing L, Yang Z, Chen L, Guo F, Fiorani F, Jackson RB, Crawford NM, Pei ZM (2004) Nitric oxide represses the Arabidopsis floral transition. Science 305:1968–1971CrossRefPubMedGoogle Scholar
  29. Hu Y, Jiang L, Wang F, Yu D (2013) Jasmonate regulates the inducer OF CBF expression-c-repeat binding factor/DRE binding factor1 cascade and freezing tolerance in Arabidopsis. Plant Cell 25:2907–2924CrossRefPubMedPubMedCentralGoogle Scholar
  30. Jackson RB, Banner JL, Jobbagy EG, Pockman WT, Wall DH (2002) Ecosystem carbon loss with woody plant invasion of grasslands. Nature 418:623–626CrossRefPubMedGoogle Scholar
  31. Jorrin-Novo JV (2009) Plant proteomics. J Proteomics 72:283–284CrossRefPubMedGoogle Scholar
  32. Kneeshaw S, Gelineau S, Tada Y, Loake GJ, Spoel SH (2014) Selective protein denitrosylation activity of thioredoxin-h5 modulates plant immunity. Mol Cell 56:153–162CrossRefPubMedGoogle Scholar
  33. Kong X, Ma L, Yang L, Chen Q, Xiang N, Yang Y, Hu X (2014) Quantitative proteomics analysis reveals that the nuclear cap-binding complex proteins Arabidopsis CBP20 and CBP80 modulate the salt stress response. J Proteome Res 13:2495–2510CrossRefPubMedGoogle Scholar
  34. Lee U, Wie C, Fernandez BO, Feelisch M, Vierling E (2008) Modulation of nitrosative stress by S-nitrosoglutathione reductase is critical for thermotolerance and plant growth in Arabidopsis. Plant Cell 20:786–802CrossRefPubMedPubMedCentralGoogle Scholar
  35. Liu L, Hausladen A, Zeng M, Que L, Heitman J, Stamler JS (2001) A metabolic enzyme for S-nitrosothiol conserved from bacteria to humans. Nature 410:490–494CrossRefPubMedGoogle Scholar
  36. Lozano-Juste J, Leon J (2011) Nitric oxide regulates DELLA content and PIF expression to promote photomorphogenesis in Arabidopsis. Plant Physiol 156:1410–1423CrossRefPubMedPubMedCentralGoogle Scholar
  37. Luo M, Liu X, Singh P, Cui Y, Zimmerli L, Wu K (2012) Chromatin modifications and remodeling in plant abiotic stress responses. Biochim Biophys Acta 1819:129–136CrossRefPubMedGoogle Scholar
  38. Mare C, Mazzucotelli E, Crosatti C, Francia E, Stanca AM, Cattivelli L (2004) Hv-WRKY38: a new transcription factor involved in cold- and drought-response in barley. Plant Mol Biol 55:399–416CrossRefPubMedGoogle Scholar
  39. Minami A, Nagao M, Ikegami K, Koshiba T, Arakawa K, Fujikawa S, Takezawa D (2005) Cold acclimation in bryophytes: low-temperature-induced freezing tolerance in Physcomitrella patens is associated with increases in expression levels of stress-related genes but not with increase in level of endogenous abscisic acid. Planta 220:414–423CrossRefPubMedGoogle Scholar
  40. Nakazawa K, Tanaka H, Arima M (1982) Rapid, simultaneous and sensitive determination of free hydroxyproline and proline in human serum by high-performance liquid chromatography. J Chromatogr 233:313–316CrossRefPubMedGoogle Scholar
  41. Neill S, Desikan R, Hancock J (2002a) Hydrogen peroxide signalling. Curr Opin Plant Biol 5:388–395CrossRefPubMedGoogle Scholar
  42. Neill SJ, Desikan R, Clarke A, Hurst RD, Hancock JT (2002b) Hydrogen peroxide and nitric oxide as signalling molecules in plants. J Exp Bot 53:1237–1247CrossRefPubMedGoogle Scholar
  43. Reichstein M, Bahn M, Mahecha MD, Kattge J, Baldocchi DD (2014) Linking plant and ecosystem functional biogeography. Proc Natl Acad Sci USA 111:13697–13702CrossRefPubMedPubMedCentralGoogle Scholar
  44. Sakamoto A, Ueda M, Morikawa H (2002) Arabidopsis glutathione-dependent formaldehyde dehydrogenase is an S-nitrosoglutathione reductase. FEBS Lett 515:20–24CrossRefPubMedGoogle Scholar
  45. Seo PJ, Kim MJ, Ryu JY, Jeong EY, Park CM (2011) Two splice variants of the IDD14 transcription factor competitively form nonfunctional heterodimers which may regulate starch metabolism. Nat Commun 2:303CrossRefPubMedGoogle Scholar
  46. Shi Y, Tian S, Hou L, Huang X, Zhang X, Guo H, Yang S (2012) Ethylene signaling negatively regulates freezing tolerance by repressing expression of CBF and type-A ARR genes in Arabidopsis. Plant Cell 24:2578–2595CrossRefPubMedPubMedCentralGoogle Scholar
  47. Spadaro D, Yun BW, Spoel SH, Chu C, Wang YQ, Loake GJ (2010) The redox switch: dynamic regulation of protein function by cysteine modifications. Physiol Plant 138:360–371CrossRefPubMedGoogle Scholar
  48. Spoel SH, Loake GJ (2011) Redox-based protein modifications: the missing link in plant immune signalling. Curr Opin Plant Biol 14:358–364CrossRefPubMedGoogle Scholar
  49. Stockinger EJ, Mao Y, Regier MK, Triezenberg SJ, Thomashow MF (2001) Transcriptional adaptor and histone acetyltransferase proteins in Arabidopsis and their interactions with CBF1, a transcriptional activator involved in cold-regulated gene expression. Nucleic Acids Res 29:1524–1533CrossRefPubMedPubMedCentralGoogle Scholar
  50. Tuskan GA, Difazio S, Jansson S, Bohlmann J, Grigoriev I, Hellsten U, Putnam N, Ralph S, Rombauts S, Salamov A et al (2006) The genome of black cottonwood, Populus trichocarpa (Torr. & Gray). Science 313:1596–1604CrossRefPubMedGoogle Scholar
  51. Vierstra RD (1996) Proteolysis in plants: mechanisms and functions. Plant Mol Biol 32:275–302CrossRefPubMedGoogle Scholar
  52. Wang L, Yang L, Yang F, Li X, Song Y, Wang X, Hu X (2010) Involvements of H2O2 and metallothionein in NO-mediated tomato tolerance to copper toxicity. J Plant Physiol 167:1298–1306CrossRefPubMedGoogle Scholar
  53. Wilson ID, Neill SJ, Hancock JT (2008) Nitric oxide synthesis and signalling in plants. Plant Cell Environ 31:622–631CrossRefPubMedGoogle Scholar
  54. Xiong L, Zhu JK (2001) Abiotic stress signal transduction in plants: molecular and genetic perspectives. Physiol Plant 112:152–166CrossRefPubMedGoogle Scholar
  55. Xiong Y, Contento AL, Nguyen PQ, Bassham DC (2007) Degradation of oxidized proteins by autophagy during oxidative stress in Arabidopsis. Plant Physiol 143:291–299CrossRefPubMedPubMedCentralGoogle Scholar
  56. Yu M, Lamattina L, Spoel SH, Loake GJ (2014) Nitric oxide function in plant biology: a redox cue in deconvolution. New Phytol 202:1142–1156CrossRefPubMedGoogle Scholar
  57. Zhao MG, Chen L, Zhang LL, Zhang WH (2009) Nitric reductase-dependent nitric oxide production is involved in cold acclimation and freezing tolerance in Arabidopsis. Plant Physiol 151:755–767CrossRefPubMedPubMedCentralGoogle Scholar
  58. Zhou QY, Tian AG, Zou HF, Xie ZM, Lei G, Huang J, Wang CM, Wang HW, Zhang JS, Chen SY (2008) Soybean WRKY-type transcription factor genes, GmWRKY13, GmWRKY21, and GmWRKY54, confer differential tolerance to abiotic stresses in transgenic Arabidopsis plants. Plant Biotechnol J 6:486–503CrossRefPubMedGoogle Scholar
  59. Zhu JH, Dong CH, Zhu JK (2007) Interplay between cold-responsive gene regulation, metabolism and RNA processing during plant cold acclimation. Curr Opin Plant Biol 10:290–295CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Tielong Cheng
    • 1
  • Jinhui Chen
    • 1
  • Abd_Allah EF
    • 2
  • Pengkai Wang
    • 1
  • Guangping Wang
    • 1
  • Xiangyang Hu
    • 3
  • Jisen Shi
    • 1
    Email author
  1. 1.Key Laboratory of Forest Genetics & Biogeography, Ministry of EducationNanjing Forest UniversityNanjingChina
  2. 2.Department of Plant Production, Faculty of Food & Agricultural SciencesKing Saud UniversityRiyadhSaudi Arabia
  3. 3.Key Laboratory for Plant Diversity and Biogeography of East Asia, Kunming Institute of BotanyChinese Academy of ScienceKunmingChina

Personalised recommendations