Skip to main content

Advertisement

SpringerLink
Log in

Identification of proteins in response to terminal drought stress in sorghum (Sorghum bicolor (L.) Moench) using two-dimensional gel-electrophoresis and MALDI-TOF-TOF MS/MS

  • Original Article
  • Published:
Indian Journal of Plant Physiology Aims and scope Submit manuscript

Abstract

Terminal drought responses in sorghum (Sorghum bicolor (L.) Moench) leaf tissue (Btx642) was investigated under post-flowering drought stress. Phenotypic data measurements and changes in protein abundance were compared between observed mean values of traits and protein spots of the well-watered (control) and drought stressed plants. Protein separation of the replicated tissue was conducted using two-dimensional gel electrophoresis. Gels were stained using Coomassie Brilliant Blue and then scanned by Molecular Imager PharosFX Plus System (BIO-RAD). Sixteen spots among 86 produced were selected based on intensity or abundance and resolution for protein identification using Matrix Assisted Laser Desorption/Ionisation—time of flight mass spectrometry (MALDI-TOF-TOF MS/MS). Nine proteins with 6 functional categories and 3 subcellular localization were identified based on MASCOT search engine of which 77.7% were significantly expressed. This study, show how plants induce intracellular signals to activate drought-response and defense pathways by up-regulating key proteins and also bypass stress condition by down regulating the rate limiting enzymes. The key metabolic pathways involved in the drought stress response were shown with 5 functional classes. The findings demonstrate novel functions of the proteins with central role in maintaining intact metabolic and photosynthetic pathways in sorghum under drought stress. Our work provides initial information as the basis for modern breeding to enhance drought tolerance and productivity in sorghum.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Abdi, A., & Asfaw, Z. (2005). In situ (on-farm) conservation dynamics and the patterns of uses of sorghum (Sorghum bicolor (L.) Moench) landraces in north Shewa and south Wollo, Central Highlands of Ethiopia. Ethiopian Journal of Biological Sciences, 4(2), 161–184.

    Google Scholar 

  • Ahmad, Y., Arya, A., Gangwar, A., Paul, S., & Bhargava, K. (2014). Proteomics in diagnosis: Past, present and future. Journal of Proteomics & Genomics, 1(1), 103.

    Google Scholar 

  • Altschul, S. F., Gish, W., Miller, W., Myers, E. W., & Lipman, D. J. (1990). Basic local alignment search tool. Journal of Molecular Biology, 215(3), 403–410.

    Article  CAS  PubMed  Google Scholar 

  • Atkin, O. K., & Macherel, D. (2009). The crucial role of plant mitochondria in orchestrating drought tolerance. Annals of Botany, 103(4), 581–597.

    Article  CAS  PubMed  Google Scholar 

  • Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72(1), 248–254.

    Article  CAS  PubMed  Google Scholar 

  • Chen, R.-D., Yu, L.-X., Greer, A. F., Cheriti, H., & Tabaeizadeh, Z. (1994). Isolation of an osmotic stress-and abscisic acid-induced gene encoding an acidic endochitinase from Lycopersicon chilense. Molecular and General Genetics MGG, 245(2), 195–202.

    Article  CAS  PubMed  Google Scholar 

  • Choudhary, J. S., Blackstock, W. P., Creasy, D. M., & Cottrell, J. S. (2001). Matching peptide mass spectra to EST and genomic DNA databases. Trends in Biotechnology, 19, 17–22.

    Article  Google Scholar 

  • Colussi, C., Albertini, M. C., Coppola, S., Rovidati, S., Galli, F., & Ghibelli, L. (2000). H2O2-induced block of glycolysis as an active ADP-ribosylation reaction protecting cells from apoptosis. The FASEB Journal, 14(14), 2266–2276. https://doi.org/10.1096/fj.00-0074com.

    Article  CAS  PubMed  Google Scholar 

  • Conesa, A., Götz, S., García-Gómez, J. M., Terol, J., Talón, M., & Robles, M. (2005). Blast2GO: A universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics, 21(18), 3674–3676.

    Article  CAS  PubMed  Google Scholar 

  • Consortium, U., et al. (2013). Update on activities at the Universal Protein Resource (UniProt) in 2013. Nucleic Acids Research, 41(D1), D43–D47.

    Article  Google Scholar 

  • Cooper, S. J., Leonard, G. A., McSweeney, S. M., Thompson, A. W., Naismith, J. H., Qamar, S., et al. (1996). The crystal structure of a class II fructose-1, 6-bisphosphate aldolase shows a novel binuclear metal-binding active site embedded in a familiar fold. Structure, 4(11), 1303–1315.

    Article  CAS  PubMed  Google Scholar 

  • Entelis, N., Brandina, I., Kamenski, P., Krasheninnikov, I. A., Martin, R. P., & Tarassov, I. (2006). A glycolytic enzyme, enolase, is recruited as a cofactor of tRNA targeting toward mitochondria in Saccharomyces cerevisiae. Genes & Development, 20(12), 1609–1620.

    Article  CAS  Google Scholar 

  • Garrels, J. I. (1989). The QUEST system for quantitative analysis of two-dimensional gels. Journal of Biological Chemistry, 264(9), 5269–5282.

    CAS  PubMed  Google Scholar 

  • Ghannoum, O. (2009). C4 photosynthesis and water stress. Annals of Botany, 103(4), 635–644.

    Article  CAS  PubMed  Google Scholar 

  • Gong, P., Zhang, J., Li, H., Yang, C., Zhang, C., Zhang, X., et al. (2010). Transcriptional profiles of drought-responsive genes in modulating transcription signal transduction, and biochemical pathways in tomato. Journal of Experimental Botany. https://doi.org/10.1093/jxb/erq167.

    Google Scholar 

  • Grabski, A. C., & Novagen, R. R. B. (2001). Preparation of protein samples for SDS-polyacrylamide gel electrophoresis: Procedures and tips. http://citeseerx.ist.psu.edu/viewdoc/summary?doi=10.1.1.378.613. Accessed 5 March 2014.

  • Haake, V., Zrenner, R., Sonnewald, U., & Stitt, M. (1998). A moderate decrease of plastid aldolase activity inhibits photosynthesis, alters the levels of sugars and starch, and inhibits growth of potato plants. The Plant Journal, 14(2), 147–157.

    Article  CAS  PubMed  Google Scholar 

  • Harris, K., Subudhi, P. K., Borrell, A., Jordan, D., Rosenow, D., Nguyen, H., et al. (2007). Sorghum stay-green QTL individually reduce post-flowering drought-induced leaf senescence. Journal of Experimental Botany, 58(2), 327–338.

    Article  CAS  PubMed  Google Scholar 

  • Hejl, A. A., Einhellig, F. A., & Rasmussen, J. A. (1993). Effects of juglone on growth, photosynthesis, and respiration. Journal of Chemical Ecology, 19(3), 559–568.

    Article  CAS  PubMed  Google Scholar 

  • Henze, K., Badr, A., Wettern, M., Cerff, R., & Martin, W. (1995). A nuclear gene of eubacterial origin in Euglena gracilis reflects cryptic endosymbioses during protist evolution. Proceedings of the National Academy of Sciences, 92(20), 9122–9126.

    Article  CAS  Google Scholar 

  • Heymans, M., & Singh, A. K. (2003). Deriving phylogenetic trees from the similarity analysis of metabolic pathways. Bioinformatics, 19(suppl 1), i138–i146.

    Article  PubMed  Google Scholar 

  • Jedmowski, C., Ashoub, A., Beckhaus, T., Berberich, T., Karas, M., & Brüggemann, W. (2014). Comparative analysis of Sorghum bicolor proteome in response to drought stress and following recovery. International Journal of Proteomics, 2014(395905), 1–10. https://doi.org/10.1155/2014/395905.

    Article  Google Scholar 

  • Jiang, S.-S., Liang, X.-N., Li, X., Wang, S.-L., Lv, D.-W., Ma, C.-Y., et al. (2012). Wheat drought-responsive grain proteome analysis by linear and nonlinear 2-DE and MALDI-TOF mass spectrometry. International Journal of Molecular Sciences, 13(12), 16065–16083.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jordan, D. R., Hunt, C. H., Cruickshank, A. W., Borrell, A. K., & Henzell, R. G. (2012). The relationship between the stay-green trait and grain yield in elite sorghum hybrids grown in a range of environments. Crop Science, 52(3), 1153–1161.

    Article  Google Scholar 

  • Kebede, H., Subudhi, P. K., Rosenow, D. T., & Nguyen, H. T. (2001). Quantitative trait loci influencing drought tolerance in grain sorghum (Sorghum bicolor L. Moench). Theoretical and Applied Genetics, 103(2–3), 266–276.

    Article  CAS  Google Scholar 

  • Kilian, J., Whitehead, D., Horak, J., Wanke, D., Weinl, S., Batistic, O., et al. (2007). The AtGenExpress global stress expression data set: Protocols, evaluation and model data analysis of UV-B light, drought and cold stress responses. The Plant Journal, 50(2), 347–363.

    Article  CAS  PubMed  Google Scholar 

  • Kim, Y.-O., Kim, J. S., & Kang, H. (2005). Cold-inducible zinc finger-containing glycine-rich RNA-binding protein contributes to the enhancement of freezing tolerance in Arabidopsis thaliana. The Plant Journal, 42(6), 890–900.

    Article  CAS  PubMed  Google Scholar 

  • Knight, H., & Knight, M. R. (2001). Abiotic stress signalling pathways: Specificity and cross-talk. Trends in Plant Science, 6(6), 262–267.

    Article  CAS  PubMed  Google Scholar 

  • Koenig, T., Menze, B. H., Kirchner, M., Monigatti, F., Parker, K. C., Patterson, T., et al. (2008). Robust prediction of the MASCOT score for an improved quality assessment in mass spectrometric proteomics. Journal of Proteome Research, 7(9), 3708–3717.

    Article  CAS  PubMed  Google Scholar 

  • Kromer, S. (1995). Respiration during photosynthesis. Annual Review of Plant Biology, 46(1), 45–70.

    Article  Google Scholar 

  • Lee, H., Guo, Y., Ohta, M., Xiong, L., Stevenson, B., & Zhu, J.-K. (2002). LOS2, a genetic locus required for cold-responsive gene transcription encodes a bi-functional enolase. The EMBO Journal, 21(11), 2692–2702.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li, Y.-H., Ou-Yang, F.-Y., Yang, C.-H., & Li, S.-Y. (2015). The coupling of glycolysis and the Rubisco-based pathway through the non-oxidative pentose phosphate pathway to achieve low carbon dioxide emission fermentation. Bioresource Technology, 187, 189–197. https://doi.org/10.1016/j.biortech.2015.03.090.

    Article  CAS  PubMed  Google Scholar 

  • Lim, H., Eng, J., Yates, J. R., III, Tollaksen, S. L., Giometti, C. S., Holden, J. F., et al. (2003). Identification of 2D-gel proteins: A comparison of MALDI/TOF peptide mass mapping to μ LC-ESI tandem mass spectrometry. Journal of the American Society for Mass Spectrometry, 14(9), 957–970. https://doi.org/10.1016/S1044-0305(03)00144-2.

    Article  CAS  PubMed  Google Scholar 

  • Liu, P., Yin, L., Deng, X., Wang, S., Tanaka, K., & Zhang, S. (2014). Aquaporin-mediated increase in root hydraulic conductance is involved in silicon-induced improved root water uptake under osmotic stress in Sorghum bicolor L. Journal of Experimental Botany, 65(17), 4747–4756.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mace, E. S., & Jordan, D. R. (2011). Integrating sorghum whole genome sequence information with a compendium of sorghum QTL studies reveals uneven distribution of QTL and of gene-rich regions with significant implications for crop improvement. Theoretical and Applied Genetics, 123(1), 169–191.

    Article  CAS  PubMed  Google Scholar 

  • Mackintosh, J. A., Veal, D. A., & Karuso, P. (2005). Fluoroprofile, a fluorescence-based assay for rapid and sensitive quantitation of proteins in solution. Proteomics, 5(18), 4673–4677. https://doi.org/10.1002/pmic.200500095.

    Article  CAS  PubMed  Google Scholar 

  • Mahalakshmi, V., & Bidinger, F. R. (2002). Evaluation of stay-green sorghum germplasm lines at ICRISAT. Crop Science, 42(3), 965–974.

    Article  Google Scholar 

  • Manners, J. M., & Casu, R. E. (2011). Transcriptome analysis and functional genomics of sugarcane. Tropical Plant Biology, 4(1), 9–21.

    Article  CAS  Google Scholar 

  • Mitchell, P. (1973). Chemiosmotic coupling in energy transduction: a logical development of biochemical knowledge. In Membrane structure and mechanisms of biological energy transduction (pp. 5–24). Springer. http://link.springer.com/chapter/10.1007/978-1-4684-2016-6_2. Accessed 20 June 2014.

  • Ndimba, B. K., Chivasa, S., Simon, W. J., & Slabas, A. R. (2005). Identification of Arabidopsis salt and osmotic stress responsive proteins using two-dimensional difference gel electrophoresis and mass spectrometry. Proteomics, 5(16), 4185–4196.

    Article  CAS  PubMed  Google Scholar 

  • Ndimba, B. K., Thomas, L. A., & Ngara, R. (2010). Sorghum 2-dimensional proteome profiles and analysis of HSP70 expression under salinity stress. Kasetsart Journal (Nat Sci), 44, 768–775.

    Google Scholar 

  • Ngara, R., Ndimba, R., Borch-Jensen, J., Jensen, O. N., & Ndimba, B. (2012). Identification and profiling of salinity stress-responsive proteins in Sorghum bicolor seedlings. Journal of Proteomics, 75(13), 4139–4150.

    Article  CAS  PubMed  Google Scholar 

  • Ogata, H., Goto, S., Sato, K., Fujibuchi, W., Bono, H., & Kanehisa, M. (1999). KEGG: Kyoto encyclopedia of genes and genomes. Nucleic Acids Research, 27(1), 29–34.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pappin, D. J., Hojrup, P., & Bleasby, A. J. (1993). Rapid identification of proteins by peptide-mass fingerprinting. Current Biology, 3(6), 327–332.

    Article  CAS  PubMed  Google Scholar 

  • Paterson, A. H., Bowers, J. E., Bruggmann, R., Dubchak, I., Grimwood, J., Gundlach, H., et al. (2009). The Sorghum bicolor genome and the diversification of grasses. Nature, 457(7229), 551–556.

    Article  CAS  PubMed  Google Scholar 

  • Pihlasalo, S., Kulmala, A., Rozwandowicz-Jansen, A., Hänninen, P., & Härmä, H. (2012). Sensitive Luminometric method for protein quantification in bacterial cell lysate based on particle adsorption and dissociation of chelated europium. Analytical Chemistry, 84(3), 1386–1393.

    Article  CAS  PubMed  Google Scholar 

  • Rabilloud, T. (2014). How to use 2D gel electrophoresis in plant proteomics. In J. V. Jorrin-Novo, S. Komatsu, W. Weckwerth, & S. Wienkoop (Eds.), Plant proteomics (pp. 43–50). Humana Press. http://dx.doi.org/10.1007/978-1-62703-631-3_4. Accessed 5 January 2017.

  • Ratnavathi, C. V., Komala, V. V., Vijaykumar, B. S., Das, I. K., & Patil, J. V. (2012). Fumonisin B1 contamination in kharik grain sorghum in India. Quality Assurance and Safety of Crops & Foods, 4(3), 146.

    Article  Google Scholar 

  • Reddy, N. R. R., Ragimasalawada, M., Sabbavarapu, M. M., Nadoor, S., & Patil, J. V. (2014). Detection and validation of stay-green QTL in post-rainy sorghum involving widely adapted cultivar, M35-1 and a popular stay-green genotype B35. BMC Genomics, 15(1), 909.

    Article  Google Scholar 

  • Sáez, D. E., & Slebe, J. C. (2000). Subcellular localization of aldolase B. Journal of Cellular Biochemistry, 78(1), 62–72.

    Article  PubMed  Google Scholar 

  • Sage, R. F. (2004). The evolution of C4 photosynthesis. New Phytologist, 161(2), 341–370.

    Article  CAS  Google Scholar 

  • Saxena, A., Tripathi, B. P., Kumar, M., & Shahi, V. K. (2009). Membrane-based techniques for the separation and purification of proteins: An overview. Advances in Colloid and Interface Science, 145(1–2), 1–22. https://doi.org/10.1016/j.cis.2008.07.004.

    Article  CAS  PubMed  Google Scholar 

  • Sayers, E. W., Barrett, T., Benson, D. A., Bolton, E., Bryant, S. H., Canese, K., et al. (2011). Database resources of the national center for biotechnology information. Nucleic Acids Research, 39(suppl 1), D38–D51.

    Article  CAS  PubMed  Google Scholar 

  • Schwender, J., Goffman, F., Ohlrogge, J. B., & Shachar-Hill, Y. (2004). Rubisco without the Calvin cycle improves the carbon efficiency of developing green seeds. Nature, 432(7018), 779–782.

    Article  CAS  PubMed  Google Scholar 

  • Shevchenko, A., Tomas, H., Havli, J., Olsen, J. V., & Mann, M. (2006). In-gel digestion for mass spectrometric characterization of proteins and proteomes. Nature Protocols, 1(6), 2856–2860.

    Article  CAS  PubMed  Google Scholar 

  • Sugihara, K., Hanagata, N., Dubinsky, Z., Baba, S., & Karube, I. (2000). Molecular characterization of cDNA encoding oxygen evolving enhancer protein 1 increased by salt treatment in the mangrove Bruguiera gymnorrhiza. Plant and Cell Physiology, 41(11), 1279–1285.

    Article  CAS  PubMed  Google Scholar 

  • Sultan, B., Roudier, P., Quirion, P., Alhassane, A., Muller, B., Dingkuhn, M., et al. (2013). Assessing climate change impacts on sorghum and millet yields in the Sudanian and Sahelian savannas of West Africa. Environmental Research Letters, 8(1), 014040.

    Article  Google Scholar 

  • Takemura, T., Hanagata, N., Sugihara, K., Baba, S., Karube, I., & Dubinsky, Z. (2000). Physiological and biochemical responses to salt stress in the mangrove, Bruguiera gymnorrhiza. Aquatic Botany, 68(1), 15–28.

    Article  CAS  Google Scholar 

  • Thiede, B., Koehler, C. J., Strozynski, M., Treumann, A., Stein, R., Zimny-Arndt, U., et al. (2013). High resolution quantitative proteomics of HeLa cells protein species using stable isotope labeling with amino acids in cell culture (SILAC), two-dimensional gel electrophoresis (2DE) and nano-liquid chromatograpohy coupled to an LTQ-OrbitrapMass spectrometer. Molecular and Cellular Proteomics, 12(2), 529–538.

    Article  CAS  PubMed  Google Scholar 

  • Walulu, R. S., Rosenow, D. T., Wester, D. B., & Nguyen, H. T. (1994). Inheritance of the stay green trait in sorghum. Crop Science, 34(4), 970–972.

    Article  Google Scholar 

  • Wang, X., Gowik, U., Tang, H., Bowers, J. E., Westhoff, P., & Paterson, A. H. (2009). Comparative genomic analysis of C4 photosynthetic pathway evolution in grasses. Genome Biology, 10(6), R68.

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang, X., Li, X., & Li, Y. (2007). A modified Coomassie Brilliant Blue staining method at nanogram sensitivity compatible with proteomic analysis. Biotechnology Letters, 29(10), 1599–1603.

    Article  CAS  PubMed  Google Scholar 

  • Woldesemayat, A. A., Van Heusden, P., Ndimba, B. K., & Christoffels, A. (2017). An integrated and comparative approach towards identification, characterization and functional annotation of candidate genes for drought tolerance in sorghum (Sorghum bicolor (L.) Moench). BMC Genetics, 18(1), 119.

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work is based on the research supported by the South African Research Chairs Initiative of the Department of Science and Technology and National Research Foundation of South Africa and the National Agricultural Proteomics Research and Service Unit (NAPRSU), University of the Western Cape. The authors thank Professor Alan Christoffels of the South African National Bioinformatics Institute (SANBI), for logistic help and kind support. The research materials were obtained from ICRISAT, India. University of South Africa provided financial and logistic support.

Author information

Authors and Affiliations

  1. South African Medical Research Council Bioinformatics Unit, South African National Bioinformatics Institute, University of the Western Cape, Private Bag X17, Belleville, 7535, South Africa

    Adugna Abdi Woldesemayat

  2. Department of Life and Consumer Sciences, College of Agriculture and Environmental Sciences, University of South Africa, Science Campus, Corner of Christiaan De Wet Road and Pioneer Avenue, Florida, Johannesburg, 1710, South Africa

    Adugna Abdi Woldesemayat

  3. Department of Agriculture and Animal Health, College of Agriculture and Environmental Sciences, University of South Africa, Science Campus, Corner of Christiaan De Wet Road and Pioneer Avenue, Florida, Johannesburg, 1710, South Africa

    David M. Modise

  4. Department of Biotechnology, University of the Western Cape, Private Bag X17, Cape Town, Western Cape, South Africa

    Bongani K. Ndimba

  5. Agricultural Research Council, Infruitech-Nietvoorbij, Private Bag X5026, Stellenbosch, 7599, South Africa

    Bongani K. Ndimba

  6. Corner of Christiaan De Wet Road and Pioneer Avenue, UNISA Science Campus, 215 NB Pityana, Florida, 1710, South Africa

    Adugna Abdi Woldesemayat

Authors
  1. Adugna Abdi Woldesemayat
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. David M. Modise
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Bongani K. Ndimba
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Adugna Abdi Woldesemayat.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

The article does not contain any studies with human or animal subjects performed by any of the authors.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Woldesemayat, A.A., Modise, D.M. & Ndimba, B.K. Identification of proteins in response to terminal drought stress in sorghum (Sorghum bicolor (L.) Moench) using two-dimensional gel-electrophoresis and MALDI-TOF-TOF MS/MS. Ind J Plant Physiol. 23, 24–39 (2018). https://doi.org/10.1007/s40502-018-0357-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s40502-018-0357-9

Keywords

Search

Navigation