Skip to main content

Quantitative Profiling of Protein Abundance and Phosphorylation State in Plant Tissues Using Tandem Mass Tags

  • Protocol
  • First Online:
Plant Proteomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2139))

Abstract

Proteins produce or regulate nearly every component of cells. Thus, the ability to quantitatively determine the protein abundance and posttranslational modification (PTM) state is a critical aspect toward our understanding of biological processes. In this chapter, we describe methods to globally quantify protein abundance and phosphorylation state using isobaric labeling with tandem mass tags followed by phosphopeptide enrichment.

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

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Wu X, Gong F, Wang W (2014) Protein extraction from plant tissues for 2DE and its application in proteomic analysis. Proteomics 14:645–658

    Article  CAS  Google Scholar 

  2. Wu X, Xiong E, Wang W et al (2014) Universal sample preparation method integrating trichloroacetic acid/acetone precipitation with phenol extraction for crop proteomic analysis. Nat Protoc 9:362–374

    Article  CAS  Google Scholar 

  3. Song G, Hsu PY, Walley JW (2018) Assessment and refinement of sample preparation methods for deep and quantitative plant proteome profiling. Proteomics 18:1800220

    Article  Google Scholar 

  4. Finkemeier I, Laxa M, Miguet L et al (2011) Proteins of diverse function and subcellular location are lysine acetylated in Arabidopsis. Plant Physiol 155:1779–1790

    Article  CAS  Google Scholar 

  5. Silva-Sanchez C, Li H, Chen S (2015) Recent advances and challenges in plant phosphoproteomics. Proteomics 15:1127–1141

    Article  CAS  Google Scholar 

  6. Rao RSP, Thelen JJ, Miernyk JA (2014) Is Lys-N(ɛ)-acetylation the next big thing in post-translational modifications? Trends Plant Sci 19:550–553

    Article  CAS  Google Scholar 

  7. Hartl M, Füßl M, Boersema PJ et al (2017) Lysine acetylome profiling uncovers novel histone deacetylase substrate proteins in Arabidopsis. Mol Syst Biol 13:949

    Article  Google Scholar 

  8. Fang X, Chen W, Zhao Y et al (2015) Global analysis of lysine acetylation in strawberry leaves. Front Plant Sci 6:739

    Article  Google Scholar 

  9. Xie X, Kang H, Liu W et al (2015) Comprehensive profiling of the rice ubiquitome reveals the significance of lysine ubiquitination in young leaves. J Proteome Res 14:2017–2025

    Article  CAS  Google Scholar 

  10. Song G, Walley JW (2016) Dynamic protein acetylation in plant–pathogen interactions. Front Plant Sci 7:421

    PubMed  PubMed Central  Google Scholar 

  11. Aguilar-Hernández V, Kim D-Y, Stankey RJ et al (2017) Mass spectrometric analyses reveal a central role for ubiquitylation in remodeling the Arabidopsis proteome during photomorphogenesis. Mol Plant 10:846–865

    Article  Google Scholar 

  12. Liu S, Yu F, Yang Z et al (2018) Establishment of dimethyl labeling-based quantitative acetylproteomics in Arabidopsis. Mol Cell Proteomics 17:1010–1027

    Article  CAS  Google Scholar 

  13. Walley JW, Shen Z, McReynolds MR et al (2018) Fungal-induced protein hyperacetylation in maize identified by acetylome profiling. Proc Natl Acad Sci 115:210–215

    Article  CAS  Google Scholar 

  14. Kelley DR (2018) E3 ubiquitin ligases: key regulators of hormone signaling in plants. Mol Cell Proteomics 17:1047–1054

    Article  CAS  Google Scholar 

  15. Pinkse MWH, Uitto PM, Hilhorst MJ et al (2004) Selective isolation at the femtomole level of phosphopeptides from proteolytic digests using 2D-NanoLC-ESI-MS/MS and titanium oxide precolumns. Anal Chem 76:3935–3943

    Article  CAS  Google Scholar 

  16. Nakagami H, Sugiyama N, Mochida K et al (2010) Large-scale comparative phosphoproteomics identifies conserved phosphorylation sites in plants. Plant Physiol 153:1161–1174

    Article  CAS  Google Scholar 

  17. Kettenbach AN, Gerber SA (2011) Rapid and reproducible single-stage phosphopeptide enrichment of complex peptide mixtures: application to general and phosphotyrosine-specific phosphoproteomics experiments. Anal Chem 83:7635–7644

    Article  CAS  Google Scholar 

  18. Marcon C, Malik WA, Walley JW et al (2015) A high-resolution tissue-specific proteome and phosphoproteome atlas of maize primary roots reveals functional gradients along the root axes. Plant Physiol 168:233–246

    Article  CAS  Google Scholar 

  19. Walley JW, Sartor RC, Shen Z et al (2016) Integration of omic networks in a developmental atlas of maize. Science 353:814–818

    Article  CAS  Google Scholar 

  20. Bantscheff M, Schirle M, Sweetman G et al (2007) Quantitative mass spectrometry in proteomics: a critical review. Anal Bioanal Chem 389:1017–1031

    Article  CAS  Google Scholar 

  21. Wiese S, Reidegeld KA, Meyer HE et al (2007) Protein labeling by iTRAQ: a new tool for quantitative mass spectrometry in proteome research. Proteomics 7:340–350

    Article  CAS  Google Scholar 

  22. Thompson A, Schäfer J, Kuhn K et al (2003) Tandem mass tags: a novel quantification strategy for comparative analysis of complex protein mixtures by MS/MS. Anal Chem 75:1895–1904

    Article  CAS  Google Scholar 

  23. Plubell DL, Wilmarth PA, Zhao Y et al (2017) Extended multiplexing of tandem mass tags (TMT) labeling reveals age and high fat diet specific proteome changes in mouse epididymal adipose tissue. Mol Cell Proteomics 16:873–890

    Article  CAS  Google Scholar 

  24. Karp NA, Huber W, Sadowski PG et al (2010) Addressing accuracy and precision issues in iTRAQ quantitation. Mol Cell Proteomics 9:1885–1897

    Article  CAS  Google Scholar 

  25. Hogrebe A, von Stechow L, Bekker-Jensen DB et al (2018) Benchmarking common quantification strategies for large-scale phosphoproteomics. Nat Commun 9:1045

    Article  Google Scholar 

  26. Song G, Brachova L, Nikolau BJ et al (2018) Heterotrimeric G-protein-dependent proteome and phosphoproteome in unstimulated Arabidopsis roots. Proteomics 18:1800323

    Article  Google Scholar 

  27. Abdulghani M, Song G, Kaur H et al (2019) Comparative analysis of the transcriptome and proteome during mouse placental development. J Proteome Res 18(5):2088–2099

    Article  CAS  Google Scholar 

  28. Mertins P, Qiao JW, Patel J et al (2013) Integrated proteomic analysis of post-translational modifications by serial enrichment. Nat Methods 10:634–637

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Justin W. Walley .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2020 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Song, G., Montes, C., Walley, J.W. (2020). Quantitative Profiling of Protein Abundance and Phosphorylation State in Plant Tissues Using Tandem Mass Tags. In: Jorrin-Novo, J., Valledor, L., Castillejo, M., Rey, MD. (eds) Plant Proteomics. Methods in Molecular Biology, vol 2139. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0528-8_11

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-0528-8_11

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0527-1

  • Online ISBN: 978-1-0716-0528-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics