Plant Molecular Biology

, Volume 83, Issue 1–2, pp 89–103 | Cite as

Proteome changes in wild and modern wheat leaves upon drought stress by two-dimensional electrophoresis and nanoLC-ESI–MS/MS

  • Hikmet BudakEmail author
  • Bala Ani Akpinar
  • Turgay Unver
  • Mine Turktas


To elucidate differentially expressed proteins and to further understand post-translational modifications of transcripts, full leaf proteome profiles of two wild emmer (Triticum turgidum ssp. dicoccoides TR39477 and TTD22) and one modern durum wheat (Triticum turgidum ssp. durum cv. Kızıltan) genotypes were compared upon 9-day drought stress using two-dimensional gel electrophoresis and nano-scale liquid chromatographic electrospray ionization tandem mass spectrometry methods. The three genotypes compared exhibit distinctive physiological responses to drought as previously shown by our group. Results demonstrated that many of the proteins were common in both wild emmer and modern wheat proteomes; of which, 75 were detected as differentially expressed proteins. Several proteins identified in all proteomes exhibited drought regulated patterns of expression. A number of proteins were observed with higher expression levels in response to drought in wild genotypes compared to their modern relative. Eleven protein spots with low peptide matches were identified as candidate unique drought responsive proteins. Of the differentially expressed proteins, four were selected and further analyzed by quantitative real-time PCR at the transcriptome level to compare with the proteomic data. The present study provides protein level differences in response to drought in modern and wild genotypes of wheat that may account for the differences of the overall responses of these genotypes to drought. Such comparative proteomics analyses may aid in the better understanding of complex drought response and may suggest candidate genes for molecular breeding studies to improve tolerance against drought stress and, thus, to enhance yields.


Drought stress Modern wheat Wild emmer nanoLC-ESI–MS/MS Proteomics 2-DE 



Authors acknowledge TUBITAK for the financial support. We would like to thank to Dr. Megan Bowman for reviewing the manuscript.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

11103_2013_24_MOESM1_ESM.xlsx (28 kb)
Supplementary material 1 (XLSX 27 kb)


  1. Abdalla KO, Rafudeen MS (2012) Analysis of the nuclear proteome of the resurrection plant Xerophyta viscosa in response to dehydration stress using iTRAQ with 2DLC and tandem mass spectrometry. J Proteomics 75(8):2361–2374. doi: 10.1016/j.jprot.2012.02.006 PubMedCrossRefGoogle Scholar
  2. Ahuja I, de Vos RC, Bones AM, Hall RD (2010) Plant molecular stress responses face climate change. Trends Plant Sci 15(12):664–674. doi: 10.1016/j.tplants.2010.08.002 PubMedCrossRefGoogle Scholar
  3. Akpinar BA, Avsar B, Lucas SJ, Budak H (2012) Plant abiotic stress signaling. Plant Signal Behav 7(11):1450–1455. doi: 10.4161/psb.21894 Google Scholar
  4. Bergman P, Edqvist J, Farbos I, Glimelius K (2000) Male-sterile tobacco displays abnormal mitochondrial atp1 transcript accumulation and reduced floral ATP/ADP ratio. Plant Mol Biol 42(3):531–544PubMedCrossRefGoogle Scholar
  5. Bhullar NK, Street K, Mackay M, Yahiaoui N, Keller B (2009) Unlocking wheat genetic resources for the molecular identification of previously undescribed functional alleles at the Pm3 resistance locus. Proc Natl Acad Sci USA 106(23):9519–9524. doi: 10.1073/pnas.0904152106 PubMedCrossRefGoogle Scholar
  6. Budak H, Kasap Z, Shearman RC, Dweikat I, Sezerman U, Mahmood A (2006) Molecular characterization of cDNA encoding resistance gene-like sequences in Buchloe dactyloides. Mol Biotechnol 34(3):293–301. doi: 10.1385/MB:34:3:293 PubMedCrossRefGoogle Scholar
  7. Caruso G, Cavaliere C, Guarino C, Gubbiotti R, Foglia P, Lagana A (2008) Identification of changes in Triticum durum L. leaf proteome in response to salt stress by two-dimensional electrophoresis and MALDI-TOF mass spectrometry. Anal Bioanal Chem 391(1):381–390. doi: 10.1007/s00216-008-2008-x Google Scholar
  8. Caruso G, Cavaliere C, Foglia P, Gubbiotti R, Samperi R, Laganà A (2009) Analysis of drought responsive proteins in wheat (Triticum durum) by 2D-PAGE and MALDI-TOF mass spectrometry. Plant Sci 177(6):570–576. doi: 10.1016/j.plantsci.2009.08.007 CrossRefGoogle Scholar
  9. Chantret N, Salse J, Sabot F, Rahman S, Bellec A, Laubin B, Dubois I, Dossat C, Sourdille P, Joudrier P, Gautier MF, Cattolico L, Beckert M, Aubourg S, Weissenbach J, Caboche M, Bernard M, Leroy P, Chalhoub B (2005) Molecular basis of evolutionary events that shaped the hardness locus in diploid and polyploid wheat species (Triticum and Aegilops). Plant Cell 17(4):1033–1045. doi: 10.1105/tpc.104.029181 PubMedCrossRefGoogle Scholar
  10. Chinnusamy V, Schumaker K, Zhu JK (2004) Molecular genetic perspectives on cross-talk and specificity in abiotic stress signalling in plants. J Exp Bot 55(395):225–236. doi: 10.1093/jxb/erh005 PubMedCrossRefGoogle Scholar
  11. Chinnusamy V, Zhu J, Zhu JK (2007) Cold stress regulation of gene expression in plants. Trends Plant Sci 12:444–451. doi: 10.1016/j.tplants.2007.07.002 Google Scholar
  12. Cruz de Carvalho MH (2008) Drought stress and reactive oxygen species: production, scavenging and signaling. Plant Signal Behav 3(3):156–165PubMedCrossRefGoogle Scholar
  13. Cui S, Huang F, Wang J, Ma X, Cheng Y, Liu J (2005) A proteomic analysis of cold stress responses in rice seedlings. Proteomics 5(12):3162–3172. doi: 10.1002/pmic.200401148 PubMedCrossRefGoogle Scholar
  14. Demirevska K, Zasheva D, Dimitrov R, Simova-Stoilova L, Stamenova M, Feller U (2009) Drought stress effects on Rubisco in wheat: changes in the Rubisco large subunit. Acta Physiol Plant 31:1129–1138. doi: 10.1007/s11738-009-0331-2 CrossRefGoogle Scholar
  15. Dixon DP, Skipsey M, Edwards R (2010) Roles for glutathione transferases in plant secondary metabolism. Phytochemistry 71(4):338–350. doi: 10.1016/j.phytochem.2009.12.012 PubMedCrossRefGoogle Scholar
  16. Ergen NZ, Budak H (2009) Sequencing over 13 000 expressed sequence tags from six subtractive cDNA libraries of wild and modern wheats following slow drought stress. Plant, Cell Environ 32(3):220–236. doi: 10.1111/j.1365-3040.2008.01915.x CrossRefGoogle Scholar
  17. Ergen NZ, Dinler G, Shearman RC, Budak H (2007) Identifying, cloning and structural analysis of differentially expressed genes upon Puccinia infection of Festuca rubra var. rubra. Gene 393(1–2):145–152PubMedCrossRefGoogle Scholar
  18. Ergen NZ, Thimmapuram J, Bohnert HJ, Budak H (2009) Transcriptome pathways unique to dehydration tolerant relatives of modern wheat. Funct Integr Genomics 9(3):377–396. doi: 10.1007/s10142-009-0123-1 PubMedCrossRefGoogle Scholar
  19. Fan P, Feng J, Jiang P, Chen X, Bao H, Nie L, Jiang D, Lv S, Kuang T, Li Y (2011) Coordination of carbon fixation and nitrogen metabolism in Salicornia europaea under salinity: comparative proteomic analysis on chloroplast proteins. Proteomics 11(22):4346–4367. doi: 10.1002/pmic.201100054 PubMedCrossRefGoogle Scholar
  20. Ferro M, Salvi D, Brugiere S, Miras S, Kowalski S, Louwagie M, Garin J, Joyard J, Rolland N (2003) Proteomics of the chloroplast envelope membranes from Arabidopsis thaliana. Mol Cell Proteomics 2(5):325–345. doi: 10.1074/mcp.M300005-MCP200 PubMedGoogle Scholar
  21. Flexas J, Medrano H (2002) Drought-inhibition of photosynthesis in C3 plants: stomatal and non-stomatal limitations revisited. Ann Bot 89(2):183–189PubMedCrossRefGoogle Scholar
  22. Galle A, Csiszar J, Secenji M, Guoth A, Cseuz L, Tari I, Gyorgyey J, Erdei L (2009) Glutathione transferase activity and expression patterns during grain filling in flag leaves of wheat genotypes differing in drought tolerance: response to water deficit. J Plant Physiol 166(17):1878–1891. doi: 10.1016/j.jplph.2009.05.016 PubMedCrossRefGoogle Scholar
  23. Gao L, Yan X, Li X, Guo G, Hu Y, Ma W, Yan Y (2011) Proteome analysis of wheat leaf under salt stress by two-dimensional difference gel electrophoresis (2D-DIGE). Phytochemistry 72(10):1180–1191. doi: 10.1016/j.phytochem.2010.12.008 PubMedCrossRefGoogle Scholar
  24. Ge P, Ma C, Wang S, Gao L, Li X, Guo G, Ma W, Yan Y (2012) Comparative proteomic analysis of grain development in two spring wheat varieties under drought stress. Anal Bioanal Chem 402(3):1297–1313. doi: 10.1007/s00216-011-5532-z PubMedCrossRefGoogle Scholar
  25. Gill SS, Tuteja N (2010) Polyamines and abiotic stress tolerance in plants. Plant Signal Behav 5(1):26–33PubMedCrossRefGoogle Scholar
  26. Groppa MD, Benavides MP (2008) Polyamines and abiotic stress: recent advances. Amino Acids 34(1):35–45. doi: 10.1007/s00726-007-0501-8 PubMedCrossRefGoogle Scholar
  27. Guo G, Ge P, Ma C, Li X, Lv D, Wang S, Ma W, Yan Y (2012) Comparative proteomic analysis of salt response proteins in seedling roots of two wheat varieties. J Proteomics 75(6):1867–1885. doi: 10.1016/j.jprot.2011.12.032 PubMedCrossRefGoogle Scholar
  28. Hrmova M, Fincher GB (2001) Structure-function relationships of beta-d-glucan endo- and exohydrolases from higher plants. Plant Mol Biol 47(1–2):73–91PubMedCrossRefGoogle Scholar
  29. Huang XY, Chao DY, Gao JP, Zhu MZ, Shi M, Lin HX (2009) A previously unknown zinc finger protein, DST, regulates drought and salt tolerance in rice via stomatal aperture control. Genes Dev 23(15):1805–1817. doi: 10.1101/gad.1812409 PubMedCrossRefGoogle Scholar
  30. Ingram J (2011) A food systems approach to researching food security and its interactions with global environmental change. Food Secur 3(4):417–431. doi: 10.1007/s12571-011-0149-9 CrossRefGoogle Scholar
  31. Kantar M, Unver T, Budak H (2010) Regulation of barley miRNAs upon dehydration stress correlated with target gene expression. Funct Integr Genomics 10(4):493–507. doi: 10.1007/s10142-010-0181-4 PubMedCrossRefGoogle Scholar
  32. Kantar M, Lucas SJ, Budak H (2011a) Drought stress: molecular genetics and genomics approaches. In: Turkan I (ed) Advances in botanical research, vol 57. Plant responses to drought and salinity stress: developments in a post-genomic era. Elsevier, Burlington, pp 445–493CrossRefGoogle Scholar
  33. Kantar M, Lucas SJ, Budak H (2011b) miRNA expression patterns of Triticum dicoccoides in response to shock drought stress. Planta 233(3):471–484. doi: 10.1007/s00425-010-1309-4 PubMedCrossRefGoogle Scholar
  34. Kausar R, Arshad M, Shahzad A, Komatsu S (2012) Proteomics analysis of sensitive and tolerant barley genotypes under drought stress. Amino Acids. doi: 10.1007/s00726-012-1338-3 PubMedGoogle Scholar
  35. Kong FJ, Oyanagi A, Komatsu S (2010) Cell wall proteome of wheat roots under flooding stress using gel-based and LC MS/MS-based proteomics approaches. Biochim Biophys Acta 1804(1):124–136. doi: 10.1016/j.bbapap.2009.09.023 PubMedCrossRefGoogle Scholar
  36. Lee H, Xiong L, Gong Z, Ishitani M, Stevenson B, Zhu J-K (2001) The Arabidopsis HOS1 gene negatively regulates cold signal transduction and encodes a RING finger protein that displays cold-regulated nucleo–cytoplasmic partitioning. Genes Dev 15:912–924. doi: 10.1101/gad.866801 Google Scholar
  37. Liu JH, Kitashiba H, Wang J, Ban Y, Moriguchi T (2007) Polyamines and their ability to provide environmental stress tolerance to plants. Plant Biotechnol J 24:117–126CrossRefGoogle Scholar
  38. Lobell DB, Schlenker W, Costa-Roberts J (2011) Climate trends and global crop production since 1980. Science 333(6042):616–620. doi: 10.1126/science.1204531 PubMedCrossRefGoogle Scholar
  39. Lucas S, Dogan E, Budak H (2011a) TMPIT1 from wild emmer wheat: first characterisation of a stress-inducible integral membrane protein. Gene 483(1–2):22–28. doi: 10.1016/j.gene.2011.05.003 PubMedCrossRefGoogle Scholar
  40. Lucas S, Durmaz E, Akpinar BA, Budak H (2011b) The drought response displayed by a DRE-binding protein from Triticum dicoccoides. Plant Physiol Biochem 49(3):346–351. doi: 10.1016/j.plaphy.2011.01.016 PubMedCrossRefGoogle Scholar
  41. Miller G, Suzuki N, Ciftci-Yilmaz S, Mittler R (2010) Reactive oxygen species homeostasis and signalling during drought and salinity stresses. Plant, Cell Environ 33(4):453–467. doi: 10.1111/j.1365-3040.2009.02041.x CrossRefGoogle Scholar
  42. Mohammadi M, Kav NN, Deyholos MK (2007) Transcriptional profiling of hexaploid wheat (Triticum aestivum L.) roots identifies novel, dehydration-responsive genes. Plant Cell Environ 30(5):630–645. doi: 10.1111/j.1365-3040.2007.01645.x Google Scholar
  43. Mohapatra PK, Patro L, Raval MK, Ramaswamy NK, Biswal UC, Biswal B (2010) Senescence-induced loss in photosynthesis enhances cell wall beta-glucosidase activity. Physiol Plant 138(3):346–355. doi: 10.1111/j.1399-3054.2009.01327.x PubMedCrossRefGoogle Scholar
  44. Moller AL, Pedas P, Andersen B, Svensson B, Schjoerring JK, Finnie C (2011) Responses of barley root and shoot proteomes to long-term nitrogen deficiency, short-term nitrogen starvation and ammonium. Plant, Cell Environ 34(12):2024–2037. doi: 10.1111/j.1365-3040.2011.02396.x CrossRefGoogle Scholar
  45. Moolna A, Bowsher CG (2010) The physiological importance of photosynthetic ferredoxin NADP + oxidoreductase (FNR) isoforms in wheat. J Exp Bot 61(10):2669–2681. doi: 10.1093/jxb/erq101 PubMedCrossRefGoogle Scholar
  46. Narita Y, Taguchi H, Nakamura T, Ueda A, Shi WM, Takabe T (2004) Characterization of the salt-inducible methionine synthase from barley leaves. Plant Sci 167(5):1009–1016. doi: 10.1016/j.plantsci.2004.05.039 CrossRefGoogle Scholar
  47. Navarre DA, Wendehenne D, Durner J, Noad R, Klessig DF (2000) Nitric oxide modulates the activity of tobacco aconitase. Plant Physiol 122(2):573–582PubMedCrossRefGoogle Scholar
  48. Neill SJ, Desikan R, Clarke A, Hurst RD, Hancock JT (2002) Hydrogen peroxide and nitric oxide as signalling molecules in plants. J Exp Bot 53(372):1237–1247PubMedCrossRefGoogle Scholar
  49. Nishizawa Y, Saruta M, Nakazono K, Nishio Z, Soma M, Yoshida T, Nakajima E, Hibi T (2003) Characterization of transgenic rice plants over-expressing the stress-inducible beta-glucanase gene Gns1. Plant Mol Biol 51(1):143–152PubMedCrossRefGoogle Scholar
  50. Nogues S, Baker NR (2000) Effects of drought on photosynthesis in Mediterranean plants grown under enhanced UV-B radiation. J Exp Bot 51(348):1309–1317PubMedCrossRefGoogle Scholar
  51. Parker R, Flowers TJ, Moore AL, Harpham NV (2006) An accurate and reproducible method for proteome profiling of the effects of salt stress in the rice leaf lamina. J Exp Bot 57(5):1109–1118. doi: 10.1093/jxb/erj134 PubMedCrossRefGoogle Scholar
  52. Peng Z, Wang M, Li F, Lv H, Li C, Xia G (2009) A proteomic study of the response to salinity and drought stress in an introgression strain of bread wheat. Mol Cell Proteomics 8(12):2676–2686. doi: 10.1074/mcp.M900052-MCP200 PubMedCrossRefGoogle Scholar
  53. Petrov VD, Van Breusegem F (2012) Hydrogen peroxide-a central hub for information flow in plant cells. AoB Plants 2012:pls014. doi: 10.1093/aobpla/pls014
  54. Qin F, Sakuma Y, Tran LSP et al (2008) Arabidopsis DREB2A-interacting proteins function as RING E3 ligases and negatively regulate plant drought stress–responsive gene expression. Plant Cell 20:1693–1707. doi: 10.1105/tpc.107.057380 Google Scholar
  55. Ravanel S, Block MA, Rippert P, Jabrin S, Curien G, Rebeille F, Douce R (2004) Methionine metabolism in plants: chloroplasts are autonomous for de novo methionine synthesis and can import S-adenosylmethionine from the cytosol. J Biol Chem 279(21):22548–22557. doi: 10.1074/jbc.M313250200 PubMedCrossRefGoogle Scholar
  56. Riccardi F, Gazeau P, de Vienne D, Zivy M (1998) Protein changes in response to progressive water deficit in maize. Quantitative variation and polypeptide identification. Plant Physiol 117(4):1253–1263PubMedCrossRefGoogle Scholar
  57. Salekdeh GH, Siopongco J, Wade LJ, Ghareyazie B, Bennett J (2002) Proteomic analysis of rice leaves during drought stress and recovery. Proteomics 2(9):1131–1145. doi: 10.1002/1615-9861(200209)2:9<1131:AID-PROT1131>3.0.CO;2-1 PubMedCrossRefGoogle Scholar
  58. Sappl PG, Heazlewood JL, Millar AH (2004) Untangling multi-gene families in plants by integrating proteomics into functional genomics. Phytochemistry 65(11):1517–1530. doi: 10.1016/j.phytochem.2004.04.021 PubMedCrossRefGoogle Scholar
  59. Shin KH, Kamal AHM, Cho K, Choi JS, Jin Y, Paek NC, Lee YW, Lee JK, Park JC, Kim HT, Heo HY, Woo SH (2011) Defense proteins are induced in wheat spikes exposed to Fusarium graminearum. Plant Omics J 4(5):270–277Google Scholar
  60. Sinclair TR (2011) Challenges in breeding for yield increase for drought. Trends Plant Sci 16(6):289–293. doi: 10.1016/j.tplants.2011.02.008 PubMedCrossRefGoogle Scholar
  61. Tanksley SD, McCouch SR (1997) Seed banks and molecular maps: unlocking genetic potential from the wild. Science 277(5329):1063–1066PubMedCrossRefGoogle Scholar
  62. Townsend DM, Manevich Y, He L, Hutchens S, Pazoles CJ, Tew KD (2009) Novel role for glutathione S-transferase pi. Regulator of protein S-Glutathionylation following oxidative and nitrosative stress. J Biol Chem 284(1):436–445. doi: 10.1074/jbc.M805586200 PubMedCrossRefGoogle Scholar
  63. Wang MC, Peng ZY, Li CL, Li F, Liu C, Xia GM (2008) Proteomic analysis on a high salt tolerance introgression strain of Triticum aestivum/Thinopyrum ponticum. Proteomics 8(7):1470–1489. doi: 10.1002/pmic.200700569 PubMedCrossRefGoogle Scholar
  64. Wingler A, Lea PJ, Quick WP, Leegood RC (2000) Photorespiration: metabolic pathways and their role in stress protection. Philos Trans R Soc Lond B Biol Sci 355(1402):1517–1529. doi: 10.1098/rstb.2000.0712 PubMedCrossRefGoogle Scholar
  65. Yan S, Tang Z, Su W, Sun W (2005) Proteomic analysis of salt stress-responsive proteins in rice root. Proteomics 5(1):235–244. doi: 10.1002/pmic.200400853 PubMedCrossRefGoogle Scholar
  66. Yoda H, Hiroi Y, Sano H (2006) Polyamine oxidase is one of the key elements for oxidative burst to induce programmed cell death in tobacco cultured cells. Plant Physiol 142(1):193–206. doi: 10.1104/pp.106.080515 PubMedCrossRefGoogle Scholar
  67. Yun SJ, Martin DJ, Gengenbach BG, Rines HW, Somers DA (1993) Sequence of a (1-3,1-4)-beta-glucanase cDNA from oat. Plant Physiol 103(1):295–296. doi: 10.1104/Pp.103.1.295 PubMedCrossRefGoogle Scholar
  68. Zhang H, Guo C, Li C, Xiao K (2008) Cloning, characterization and expression analysis of two superoxide dismutase (SOD) genes in wheat (Triticum aestivum L.). Frontiers Agric China 2(2):141–149. doi: 10.1007/s11703-008-0023-5 CrossRefGoogle Scholar
  69. Zhang M, Li G, Huang W, Bi T, Chen G, Tang Z, Su W, Sun W (2010) Proteomic study of Carissa spinarum in response to combined heat and drought stress. Proteomics 10(17):3117–3129. doi: 10.1002/pmic.200900637 PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Hikmet Budak
    • 1
    Email author
  • Bala Ani Akpinar
    • 1
  • Turgay Unver
    • 1
  • Mine Turktas
    • 1
  1. 1.Biological Sciences and Bioengineering Program, Faculty of Engineering and Natural SciencesSabancı UniversityTuzla, IstanbulTurkey

Personalised recommendations