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Isolation and Analysis of Cell Wall Proteome in Elsholtzia splendens Roots Using ITRAQ with LC–ESI–MS/MS

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Abstract

Cell wall proteins (CWPs) are a prime site for signal perception and defense responses to environmental stresses. To gain further insights into CWPs and their molecular function, traditional techniques (e.g., two-dimensional gel electrophoresis) may be ineffective for special proteins. Elsholtzia splendens is a copper-tolerant plant species that grow on copper deposits. In this study, a fourplex isobaric tag was used for relative and absolute quantitation with liquid chromatography–tandem mass spectrometry approach to analyze the root CWPs of E. splendens. A total of 479 unique proteins were identified, including 121 novel proteins. Approximately 80.79 % of the proteins were extracted in the CaCl2 fraction, 16.08 % were detected in the NaCl fraction, and 3.13 % were identified in both fractions. The identified proteins have been involved in various processes, including cell wall remodeling, signal transduction, defense, and carbohydrate metabolism, thereby indicating a complex regulatory network in the apoplast of E. splendens roots. This study presents the first large-scale analysis of CWPs in metal-tolerant plants, which may be of paramount importance to understand the molecular functions and metabolic pathways in the root cell wall of copper-tolerant plants.

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References

  1. Jarvis, M. C. (2009). Plant cell walls: supramolecular assembly, signalling and stress. Structural Chemistry, 20(2), 245–253.

    Article  CAS  Google Scholar 

  2. Krzesłowska, M. (2011). The cell wall in plant cell response to trace metals: polysaccharide remodeling and its role in defense strategy. Acta Physiologiae Plantarum, 33(1), 35–51.

    Article  Google Scholar 

  3. Zhu, J., Chen, S., Alvarez, S., Asirvatham, V. S., Schachtman, D. P., Wu, Y., & Sharp, R. E. (2006). Cell wall proteome in the maize primary root elongation zone. I. Extraction and identification of water-soluble and lightly ionically bound proteins. Plant Physiology, 140(1), 311–325.

    Article  CAS  Google Scholar 

  4. Jamet, E., Albenne, C., Boudart, G., Irshad, M., Canut, H., & Pont‐Lezica, R. (2008). Recent advances in plant cell wall proteomics. Proteomics, 8(4), 893–908.

    Article  CAS  Google Scholar 

  5. Held, M. A., Tan, L., Kamyab, A., Hare, M., Shpak, E., & Kieliszewski, M. J. (2004). Di-isodityrosine is the intermolecular cross-link of isodityrosine-rich extensin analogs cross-linked in vitro. Journal of Biological Chemistry, 279(53), 55474–55482.

    Article  CAS  Google Scholar 

  6. Boudart, G., Jamet, E., Rossignol, M., Lafitte, C., Borderies, G., Jauneau, A., Esquerré‐Tugayé, M. T., & Pont‐Lezica, R. (2005). Cell wall proteins in apoplastic fluids of Arabidopsis thaliana rosettes: identification by mass spectrometry and bioinformatics. Proteomics, 5(1), 212–221.

    Article  CAS  Google Scholar 

  7. Jamet, E., Boudart, G., Borderies, G., Charmont, S., Lafitte, C., Rossignol, M., Canut, H., & Pont-Lezica, R. (2008). In methods in molecular biology. vol. 2: 2D PAGE: Sample Preparation and Fractionation, Humana, Munich, Germany, pp. 187-201.

  8. Dani, V., Simon, W. J., Duranti, M., & Croy, R. R. (2005). Changes in the tobacco leaf apoplast proteome in response to salt stress. Proteomics, 5(3), 737–745.

    Article  CAS  Google Scholar 

  9. Soares, N. C., Francisco, R., Vielba, J. M., Ricardo, C. P., & Jackson, P. A. (2009). Associating wound-related changes in the apoplast proteome of Medicago with early steps in the ROS signal-transduction pathway. Journal of Proteome Research, 8(5), 2298–2309.

    Article  CAS  Google Scholar 

  10. Jamet, E., Canut, H., Boudart, G., & Pont-Lezica, R. F. (2006). Cell wall proteins: a new insight through proteomics. Trends in Plant Science, 11(1), 33–39.

    Article  CAS  Google Scholar 

  11. Bhushan, D., Pandey, A., Choudhary, M. K., Datta, A., Chakraborty, S., & Chakraborty, N. (2007). Comparative proteomics analysis of differentially expressed proteins in chickpea extracellular matrix during dehydration stress. Molecular & Cellular Proteomics, 6(11), 1868–1884.

    Article  CAS  Google Scholar 

  12. Pandey, A., Rajamani, U., Verma, J., Subba, P., Chakraborty, N., Datta, A., Chakraborty, S., & Chakraborty, N. (2010). Identification of extracellular matrix proteins of rice (Oryza sativa L.) involved in dehydration-responsive network: a proteomic approach. Journal Proteome Research, 9(7), 3443–3464.

    Article  CAS  Google Scholar 

  13. Floerl, S., Druebert, C., Majcherczyk, A., Karlovsky, P., Kües, U., & Polle, A. (2008). Defence reactions in the apoplastic proteome of oilseed rape (Brassica napus var. napus) attenuate Verticillium longisporum growth but not disease symptoms. BMC Plant Biology, 8(1), 129.

    Article  Google Scholar 

  14. Goulet, C., Goulet, C., Goulet, M. C., & Michaud, D. (2010). 2‐DE proteome maps for the leaf apoplast of Nicotiana benthamiana. Proteomics, 10(13), 2536–2544.

    Article  CAS  Google Scholar 

  15. Irshad, M., Canut, H., Borderies, G., Pont-Lezica, R., & Jamet, E. (2008). A new picture of cell wall protein dynamics in elongating cells of Arabidopsis thaliana: confirmed actors and newcomers. BMC Plant Biology, 8(1), 94.

    Article  Google Scholar 

  16. Chivasa, S., Ndimba, B. K., Simon, W. J., Robertson, D., Yu, X. L., Knox, J. P., Bolwell, P., & Slabas, A. R. (2002). Proteomic analysis of the Arabidopsis thaliana cell wall. Electrophoresis, 23(11), 1754–1765.

    Article  CAS  Google Scholar 

  17. Borderies, G., Jamet, E., Lafitte, C., Rossignol, M., Jauneau, A., Boudart, G., Monsarrat, B., Esquerré‐Tugayé, M. T., Boudet, A., & Pont‐Lezica, R. (2003). Proteomics of loosely bound cell wall proteins of Arabidopsis thaliana cell suspension cultures: a critical analysis. Electrophoresis, 24(19‐20), 3421–3432.

    Article  CAS  Google Scholar 

  18. Albenne, C., Canut, H., & Jamet, E. (2013). Plant cell wall proteomics: the leadership of Arabidopsis thaliana. Frontiers in Plant Science, 4, 1–13.

    Article  Google Scholar 

  19. Valente, M. A. S., Faria, J. A., Soares-Ramos, J. R., Reis, P. A., Pinheiro, G. L., Piovesan, N. D., Morais, A. T., Menezes, C. C., Cano, M. A., & Fietto, L. G. (2009). The ER luminal binding protein (BiP) mediates an increase in drought tolerance in soybean and delays drought-induced leaf senescence in soybean and tobacco. Journal of Experimental Botany, 60(2), 533–546.

    Article  CAS  Google Scholar 

  20. Chen, X. Y., Kim, S. T., Cho, W. K., Rim, Y., Kim, S., Kim, S. W., Kang, K. Y., Park, Z. Y., & Kim, J. Y. (2009). Proteomics of weakly bound cell wall proteins in rice calli. Journal of Plant Physiology, 166(7), 675–685.

    Article  CAS  Google Scholar 

  21. Millar, D. J., Whitelegge, J. P., Bindschedler, L. V., Rayon, C., Boudet, A. M., Rossignol, M., Borderies, G., & Bolwell, G. P. (2009). The cell wall and secretory proteome of a tobacco cell line synthesising secondary wall. Proteomics, 9(9), 2355–2372.

    Article  CAS  Google Scholar 

  22. Bhushan, D., Pandey, A., Chattopadhyay, A., Choudhary, M. K., Chakraborty, S., Datta, A., & Chakraborty, N. (2006). Extracellular matrix proteome of chickpea (Cicer a rietinum L.) illustrates pathway abundance, novel protein functions and evolutionary perspect. Journal of Proteome Research, 5(7), 1711–1720.

    Article  CAS  Google Scholar 

  23. Witzel, K., Shahzad, M., Matros, A., Mock, H. P., & Mühling, K. H. (2011). Comparative evaluation of extraction methods for apoplastic proteins from maize leaves. Plant Methods, 7(1), 48–52.

    Article  CAS  Google Scholar 

  24. Watson, B. S., Lei, Z., Dixon, R. A., & Sumner, L. W. (2004). Proteomics of Medicago sativa cell walls. Phytochemistry, 65(12), 1709–1720.

    Article  CAS  Google Scholar 

  25. Soares, N. C., Francisco, R., Ricardo, C. P., & Jackson, P. A. (2007). Proteomics of ionically bound and soluble extracellular proteins in Medicago truncatula leaves. Proteomics, 7(12), 2070–2082.

    Article  CAS  Google Scholar 

  26. Verdonk, J. C., Hatfield, R. D., & Sullivan, M. L. (2012). Proteomic analysis of cell walls of two developmental stages of alfalfa stems. Frontiers in Plant Science, 3, 1–13.

    Article  Google Scholar 

  27. Day, A., Fénart, S., Neutelings, G., Hawkins, S., Rolando, C., & Tokarski, C. (2013). Identification of cell wall proteins in the flax (Linum usitatissimum) stem. Proteomics, 13(5), 812–825.

    Article  CAS  Google Scholar 

  28. Wang, S. B., Hu, Q., Sommerfeld, M., & Chen, F. (2004). Cell wall proteomics of the green alga Haematococcus pluvialis (Chlorophyceae). Proteomics, 4(3), 692–708.

    Article  CAS  Google Scholar 

  29. Frigeri, L. G., Radabaugh, T. R., Haynes, P. A., & Hildebrand, M. (2013). Identification of proteins from a cell wall fraction of the diatom thalassiosira pseudonana insights into silica structure formation. Molecular & Cellular Proteomics 2006, 5(1), 182–193.

    Article  Google Scholar 

  30. Wu, W. W., Wang, G., Baek, S. J., & Shen, R. F. (2006). Comparative study of three proteomic quantitative methods, DIGE, cICAT, and iTRAQ, using 2D gel-or LC-MALDI TOF/TOF. Journal of Proteome Research, 5(3), 651–658.

    Article  CAS  Google Scholar 

  31. Yang, M. J., Yang, X. E., & Römheld, V. (2002). Growth and nutrient composition of Elsholtzia splendens Nakai under copper toxicity. Journal of Plant Nutrition, 25(7), 1359–1375.

    Article  CAS  Google Scholar 

  32. Lou, L., Shen, Z., & Li, X. (2004). The copper tolerance mechanisms of Elsholtzia haichowensis, a plant from copper-enriched soils. Environmental and Experimental Botany, 51(2), 111–120.

    Article  CAS  Google Scholar 

  33. Harrison, S., Lepp, N., & Phipps, D. (1979). Uptake of Copper by excised roots II. Copper desorption from the free space. Zeitschrift für Pflanzenphysiologie, 94(1), 27–34.

    Article  CAS  Google Scholar 

  34. Feiz, L., Irshad, M., Pont-Lezica, R. F., Canut, H., & Jamet, E. (2006). Evaluation of cell wall preparations for proteomics: a new procedure for purifying cell walls from Arabidopsis hypocotyls. Plant Methods, 2, 10.

    Article  Google Scholar 

  35. 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  Google Scholar 

  36. Honjoh, K. I., Mimura, A., Kuroiwa, E., Hagisako, T., Suga, K., Shimizu, H., Dubeu, R. S., Miyamoto, T., Hatano, S., & IIo, M. (2003). Purification and characterization of two isoforms of glucose 6-phosphate dehydrogenase (G6PDH) from Chlorella vulgaris C-27. Bioscience, Biotechnology, and Biochemistry, 67(9), 1888–1896.

    Article  CAS  Google Scholar 

  37. Ye, J., Fang, L., Zheng, H., Zhang, Y., Chen, J., Zhang, Z., Wang, J., Li, S., Li, R., & Bolund, L. (2006). WEGO: a web tool for plotting GO annotations. Nucleic Acids Research, 34(suppl 2), W293–W297.

    Article  CAS  Google Scholar 

  38. Kanehisa, M., & Goto, S. (2000). KEGG: kyoto encyclopedia of genes and genomes. Nucleic Acids Research, 28(1), 27–30.

    Article  CAS  Google Scholar 

  39. Feiz, L., Irshad, M., Pont-Lezica, R. F., Canut, H., & Jamet, E. (2006). Evaluation of cell wall preparations for proteomics: a new procedure for purifying cell walls from Arabidopsis hypocotyls. Plant Methods, 2(1), 10.

    Article  Google Scholar 

  40. Kwon, H. K., Yokoyama, R., & Nishitani, K. (2005). A proteomic approach to apoplastic proteins involved in cell wall regeneration in protoplasts of Arabidopsis suspension-cultured cells. Plant and Cell Physiology, 46(6), 843–857.

    Article  CAS  Google Scholar 

  41. Xu, Z., Escamilla-Treviño, L., Zeng, L., Lalgondar, M., Bevan, D., Winkel, B., Mohamed, A., Cheng, C. L., Shih, M. C., & Poulton, J. (2004). Functional genomic analysis of Arabidopsis thaliana glycoside hydrolase family 1. Plant Molecular Biology, 55(3), 343–367.

    Article  CAS  Google Scholar 

  42. Narula, K., Elagamey, E., Datta, A., Chakraborty, N., Chakraborty, S., & Goyal, A. (2012). Comparative analyses of extracellular matrix proteome: an under-explored area in plant research. Crop Plants 7, 145-166.

  43. Derbyshire, P., Findlay, K., McCann, M. C., & Roberts, K. (2007). Cell elongation in Arabidopsis hypocotyls involves dynamic changes in cell wall thickness. Journal of Experimental Botany, 58(8), 2079–2089.

    Article  CAS  Google Scholar 

  44. Kevil, C. G., De Benedetti, A., Payne, D. K., Coe, L. L., Laroux, F. S., & Alexander, J. S. (1996). Translational regulation of vascular permeability factor by eukaryotic initiation factor 4E: implications for tumor angiogenesis. International Journal of Cancer, 65(6), 785–790.

    Article  CAS  Google Scholar 

  45. Lu, P., Vogel, C., Wang, R., Yao, X., & Marcotte, E. M. (2006). Absolute protein expression profiling estimates the relative contributions of transcriptional and translational regulation. Nature Biotechnology, 25(1), 117–124.

    Article  Google Scholar 

  46. Naujokat, C., & Hoffmann, S. (2002). Role and function of the 26S proteasome in proliferation and apoptosis. Laboratory Investigation, 82(8), 965–980.

    Article  CAS  Google Scholar 

  47. Eikmanns, B. J., Thum-Schmitz, N., Eggeling, L., Lüdtke, K. U., & Sahm, H. (2013). Nucleotide sequence, expression and transcriptional analysis of the Corynebacterium glutamicum gltA gene encoding citrate synthase. Microbiology, 1994, 140(8), 1817–1828.

  48. Jackson, R. J., Hellen, C., & Pestova, T. V. (2010). The mechanism of eukaryotic translation initiation and principles of its regulation. Nature Reviews Molecular Cell Biology, 11(2), 113–127.

    Article  CAS  Google Scholar 

  49. Hor, L., Dobson, R. C., Downton, M. T., Wagner, J., Hutton, C. A., & Perugini, M. A. (2013). Dimerization of bacterial diaminopimelate epimerase is essential for catalysis. Journal of Biological Chemistry, 288(13), 9238–9248.

    Article  CAS  Google Scholar 

  50. Xu, S., Butkevich, A. N., Yamada, R., Zhou, Y., Debnath, B., Duncan, R., Zandi, E., Petasis, N. A., & Neamati, N. (2012). Discovery of an orally active small-molecule irreversible inhibitor of protein disulfide isomerase for ovarian cancer treatment. Proceedings of the National Academy of Sciences, 109(40), 16348–16353.

    Article  CAS  Google Scholar 

  51. Richard, J. P. (2012). A paradigm for enzyme-catalyzed proton transfer at carbon: triosephosphate isomerase. Biochemistry, 51(13), 2652–2661.

    Article  CAS  Google Scholar 

  52. Chivasa, S., Ndimba, B. K., Simon, W. J., Lindsey, K., & Slabas, A. R. (2005). Extracellular ATP functions as an endogenous external metabolite regulating plant cell viability. The Plant Cell Online, 17(11), 3019–3034.

    Article  CAS  Google Scholar 

  53. Ndimba, B. K., Chivasa, S., Hamilton, J. M., Simon, W. J., & Slabas, A. R. (2003). Proteomic analysis of changes in the extracellular matrix of Arabidopsis cell suspension cultures induced by fungal elicitors. Proteomics, 3(6), 1047–1059.

    Article  CAS  Google Scholar 

  54. Stock, J., Ninfa, A., & Stock, A. (1989). Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiological Reviews, 53(4), 450.

    CAS  Google Scholar 

  55. Chivasa, S., Simon, W. J., Yu, X. L., Yalpani, N., & Slabas, A. R. (2005). Pathogen elicitor‐induced changes in the maize extracellular matrix proteome. Proteomics, 5(18), 4894–4904.

    Article  CAS  Google Scholar 

  56. Kasai, H., & Aosaki, T. (1989). Modulation of Ca-channel current by an adenosine analog mediated by a GTP-binding protein in chick sensory neurons. Pflügers Archiv, 414(2), 145–149.

    Article  CAS  Google Scholar 

  57. Schwacke, R., Schneider, A., Van der Graaff, E., Fischer, K., Catoni, E., Desimone, M., Frommer, W. B., Flügge, U. I., & Kunze, R. (2003). ARAMEMNON, a novel database for Arabidopsis integral membrane proteins. Plant Physiology, 131(1), 16–26.

    Article  CAS  Google Scholar 

  58. Murti, K., Smith, H. T., & Fried, V. A. (1988). Ubiquitin is a component of the microtubule network. Proceedings of the National Academy of Sciences, 85(9), 3019–3023.

    Article  CAS  Google Scholar 

  59. Gokulakannan, G. G., & Niehaus, K. (2010). Characterization of the Medicago truncatula cell wall proteome in cell suspension culture upon elicitation and suppression of plant defense. Journal of Plant Physiology, 167(18), 1533–1541.

    Article  CAS  Google Scholar 

  60. Passardi, F., Penel, C., & Dunand, C. (2004). Performing the paradoxical: how plant peroxidases modify the cell wall. Trends in Plant Science, 9(11), 534–540.

    Article  CAS  Google Scholar 

  61. Higuchi, M., Pischke, M. S., Mähönen, A. P., Miyawaki, K., Hashimoto, Y., Seki, M., Kobayashi, M., Shinozaki, K., Kato, T., & Tabata, S. (2004). In planta functions of the Arabidopsis cytokinin receptor family. Proceedings of the National Academy of Sciences of the United States of America, 101(23), 8821–8826.

    Article  CAS  Google Scholar 

  62. Imin, N., Kerim, T., Rolfe, B. G., & Weinman, J. J. (2004). Effect of early cold stress on the maturation of rice anthers. Proteomics, 4(7), 1873–1882.

    Article  CAS  Google Scholar 

  63. Zhang, H., Xia, Y., Wang, G., & Shen, Z. (2008). Excess copper induces accumulation of hydrogen peroxide and increases lipid peroxidation and total activity of copper–zinc superoxide dismutase in roots of Elsholtzia haichowensis. Planta, 227(2), 465–475.

    Article  CAS  Google Scholar 

  64. Timperio, A. M., Egidi, M. G., & Zolla, L. (2008). Proteomics applied on plant abiotic stresses: role of heat shock proteins (HSP). Journal of Proteomics, 71(4), 391–411.

    Article  CAS  Google Scholar 

  65. Abe, S., Sakai, M., Yagi, K., Hagino, T., Ochi, K., Shibata, K., & Davies, E. (2003). A Tudor protein with multiple SNc domains from pea seedlings: cellular localization, partial characterization, sequence analysis, and phylogenetic relationships. Journal of Experimental Botany, 54(384), 971–983.

    Article  CAS  Google Scholar 

  66. Qu, Z. L., Zhong, N. Q., Wang, H. Y., Chen, A. P., Jian, G. L., & Xia, G. X. (2006). Ectopic expression of the cotton non-symbiotic hemoglobin gene GhHbd1 triggers defense responses and increases disease tolerance in Arabidopsis. Plant and Cell Physiology, 47(8), 1058–1068.

    Article  CAS  Google Scholar 

  67. Doyle, V., Virji, S., & Crompton, M. (1999). Evidence that cyclophilin-A protects cells against oxidative stress. Biochemistry Journal, 341, 127–132.

    Article  CAS  Google Scholar 

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Acknowledgments

This study was supported by the National Natural Science Foundation of China (41422107, 21177109), the Fundamental Research Funds for the Central Universities (2015XZZX004-15), and Zhejiang University K. P. Chao’s High Technology Development Foundation (2014RC011). The authors would like to express their deepest gratitude to Dongfeng Liu from the College of Agriculture and Biotechnology in Zhejiang University for her kind help during the experiments.

Conflicts of Interest

The authors declare no conflict of interest.

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Authors and Affiliations

  1. Institute of Environmental Science and Technology, College of Environmental and Resource Sciences, Zhejiang University, Hangzhou, 310058, People’s Republic of China

    Tingting Liu, Chaofeng Shen & Jiyan Shi

  2. Institute of Environmental Engineering, Tongji Zhejiang College, Jiaxing, 314051, People’s Republic of China

    Tingting Liu

  3. Wenzhou environmental monitoring center, Wenzhou, 325000, People’s Republic of China

    Canke Huang

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  1. Tingting Liu
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  4. Jiyan Shi
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Correspondence to Jiyan Shi.

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Highlights

• First large-scale analysis of CWPs in metal-tolerant plants

• Enhancement of knowledge regarding complexity of apoplast proteins in metal-tolerant plants

• Clues for further functional research of target proteins associated with metal-tolerant plants

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Fig. S1

Workflow of the experimental design for iTRAQ labeling showing biological replicates. Cell wall proteins from CaCl2 extraction were labeled with 114 and 116, respectively; Cell wall proteins from NaCl extraction were labeled with 115 and 117, respectively (DOC 28 kb)

Table S1

Subcellular localization and GO terms corresponding to the biological process of the 479 root cell wall proteins (XLS 506 kb)

Table S2

Relative quantitative information in CaCl2- and NaCl-extracted protein samples using 4-plex iTRAQ technology (XLS 277 kb)

Table S3

Peptides from 479 unique proteins (XLS 1841 kb)

Table S4

A total of 479 CWPs were identified including 121 novel proteins (DOC 774 kb)

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Liu, T., Huang, C., Shen, C. et al. Isolation and Analysis of Cell Wall Proteome in Elsholtzia splendens Roots Using ITRAQ with LC–ESI–MS/MS. Appl Biochem Biotechnol 176, 1174–1194 (2015). https://doi.org/10.1007/s12010-015-1638-7

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