Using Functional Proteome Microarrays to Study Protein Lysine Acetylation

  • Jin-ying Lu
  • Yu-yi Lin
  • Jef D. Boeke
  • Heng Zhu
Part of the Methods in Molecular Biology book series (MIMB, volume 981)


Emergence of proteome microarray provides a versatile platform to globally explore biological functions of broad significance. In the past decade, researchers have successfully fabricated functional proteome microarrays by printing individually purified proteins at a high-throughput, proteome-wide scale on one single slide. These arrays have been used to profile protein posttranslational modifications, including phosphorylation, ubiquitylation, acetylation, and nitrosylation. In this chapter, we summarize our work of using the yeast proteome microarrays to connect protein lysine acetylation substrates to their upstream modifying enzyme, the nucleosome acetyltransferase of H4 (NuA4), which is the only essential acetyltransferase in yeast. We further prove that the reversible acetylation on critical cell metabolism-related enzymes controls life span in yeast. Our studies represent a paradigm shift for the functional dissection of a crucial acetylation enzyme affecting aging and longevity pathways.

Key words

Protein chip Posttranslational modification Lysine acetylation Functional proteome microarray Aging Longevity 


  1. 1.
    Smith MG, Jona G, Ptacek J, Devgan G, Zhu H, Zhu X, Snyder M (2005) Global analysis of protein function using protein microarrays. Mech Ageing Dev 126:171–175PubMedCrossRefGoogle Scholar
  2. 2.
    Chen C, Zhu H (2006) Protein microarrays. Biotechniques 40:432–439CrossRefGoogle Scholar
  3. 3.
    Tao SC, Chen CS, Zhu H (2007) Applications of protein microarray technology. Comb Chem High Throughput Screen 10:706–718PubMedCrossRefGoogle Scholar
  4. 4.
    Zhu H, Bilgin M, Bangham R, Hall D, Casamayor A, Bertone P, Lan N, Jansen R, Bidlingmaier S, Houfek T, Mitchell T, Miller P, Dean RA, Gerstein M, Snyder M (2001) Global analysis of protein activities using proteome chips. Science 293:2101–2105PubMedCrossRefGoogle Scholar
  5. 5.
    Hall DA, Zhu H, Zhu X, Royce T, Gerstein M, Snyder M (2004) Regulation of gene expression by a metabolic enzyme. Science 306: 482–484PubMedCrossRefGoogle Scholar
  6. 6.
    Ho SW, Jona G, Chen CT, Johnston M, Snyder M (2006) Linking DNA-binding proteins to their recognition sequences by using protein microarrays. Pro Natl Acad Sci USA 103(26): 9940–9945CrossRefGoogle Scholar
  7. 7.
    Hu S, Xie Z, Onishi A, Yu X, Jiang L, Lin J, Rho HS, Woodard C, Wang H, Jeong JS, Long S, He X, Wade H, Blackshaw S, Qian J, Zhu H (2009) Profiling the human protein–DNA interactome reveals ERK2 as a transcriptional repressor of interferon signalling. Cell 139(3):610–622PubMedCrossRefGoogle Scholar
  8. 8.
    Zhu J, Gopinath K, Murali A, Yi G, Hayward SD, Zhu H, Kao C (2007) RNA binding proteins that inhibit RNA virus infection. Proc Natl Acad Sci USA 104:3129–3134PubMedCrossRefGoogle Scholar
  9. 9.
    Huang J, Zhu H, Haggarty SJ, Spring DR, Hwang H, Jin F, Snyder M, Schreiber SL (2004) Finding new components of the target of rapamycin (TOR) signaling network through chemical genetics and proteome chips. Proc Natl Acad Sci USA 101:16594–16599PubMedCrossRefGoogle Scholar
  10. 10.
    Zhu H, Klemic JF, Chang S, Bertone P, Casamayor A, Klemic KG, Smith D, Gerstein M, Reed MA, Snyder M (2000) Analysis of yeast protein kinases using protein chips. Nat Genet 26:283–289PubMedCrossRefGoogle Scholar
  11. 11.
    Kafadar KA, Zhu H, Snyder M, Cyert MS (2003) Negative regulation of calcineurin signaling by Hrr25p, a yeast homolog of casein kinase I. Genes Dev 17:2698–2708PubMedCrossRefGoogle Scholar
  12. 12.
    Ptacek J, Devgan G, Michaud G, Zhu H et al (2005) Global analysis of protein phosphorylation in yeast. Nature 438:679–684PubMedCrossRefGoogle Scholar
  13. 13.
    Tao SC, Li Y, Zhou J, Qian J, Schnaar RL, Zhang Y, Goldstein IJ, Zhu H, Schneck JP (2008) Lectin microarrays identify cell-specific and functionally significant cell surface glycan markers. Glycobiology 18:761–769PubMedCrossRefGoogle Scholar
  14. 14.
    Zhu J, Liao G, Shan L, Zhang J, Chen MR, Hayward GS, Hayward SD, Desai P, Zhu H (2009) Protein array identification of substrates of the Epstein-Barr Virus protein kinase BGLF4. J Virol 83:5219–5231PubMedCrossRefGoogle Scholar
  15. 15.
    Kung L, Tao SC, Qian J, Smith M, Snyder M, Zhu H (2009) Global analysis of the glycoproteome in S. cerevisiae reveals new roles for protein glycosylation. Mol Syst Biol 5:308PubMedCrossRefGoogle Scholar
  16. 16.
    Lu JY, Lin YY, Tao SC, Zhu J, Pickart CM, Qian J, Zhu H (2008) Functional dissection of a HECT ubiquitin E3 ligase. Mol Cell Proteomics 7:35–45PubMedGoogle Scholar
  17. 17.
    Lin YY, Lu JY, Zhang J, Walter W, Dang W, Wan J, Tao SC, Qian J, Zhao Y, Boeke JD, Berger SL, Zhu H (2009) Protein acetylation microarray reveals NuA4 controls key metabolic target regulating gluconeogenesis. Cell 136:1073–1084PubMedCrossRefGoogle Scholar
  18. 18.
    Thao S, Chen CS, Zhu H, Escalante-Semerena JC (2010) Nε-lysine acetylation of a bacterial transcription factor inhibits Its DNA-binding activity. PLoS One 5(12):15123CrossRefGoogle Scholar
  19. 19.
    Lu JY, Lin YY, Sheu JC, Wu JT, Lee FJ, Chen Y, Lin MI, Chiang FT, Tai TY, Berger SL, Zhao Y, Tsai KS, Zhu H, Chuang LM, Boeke JD (2011) Acetylation of AMPK controls intrinsic aging independently of caloric restriction. Cell 146:969–979PubMedCrossRefGoogle Scholar
  20. 20.
    Oh YH, Hong MY, Jin Z, Lee T, Han MK, Park S, Kim HS (2007) Chip-based analysis of SUMO (small ubiquitin-like modifier) conjugation to a target protein. Biosens Bioelectron 22(7):1260–1267PubMedCrossRefGoogle Scholar
  21. 21.
    Del Rincón SV, Rogers J, Widschwendter M, Sun D, Sieburg HB, Spruck C (2010) Development and validation of a method for profiling post-translational modification activities using protein microarrays. PLoS One 5(6): e11332PubMedCrossRefGoogle Scholar
  22. 22.
    Sterner DE, Berger SL (2000) Acetylation of histones and transcription-related factors. Microbiol Mol Biol Rev 64:435PubMedCrossRefGoogle Scholar
  23. 23.
    Smith ER, Eisen A, Gu W, Sattah M, Pannuti A, Zhou J, Cook RG, Lucchesi JC, Allis CD (1998) ESA1 is a histone acetyltransferase that is essential for growth in yeast. Proc Natl Acad Sci USA 95:3561–3565PubMedCrossRefGoogle Scholar
  24. 24.
    Li Y, Yokota T, Gama V, Yoshida T, Gomez JA, Ishikawa K, Sasaguri H, Cohen HY, Sinclair DA, Mizusawa H, Matsuyama S (2007) Bax-inhibiting peptide protects cells from polyglutamine toxicity caused by Ku70 acetylation. Cell Death Differ 14:2058–2067PubMedCrossRefGoogle Scholar
  25. 25.
    Burlini N, Lamponi S, Radrizzani M, Monti E, Tortora P (1987) Identification of a phosphorylated form of phosphoenolpyruvate carboxykinase from the yeast Saccharomyces cerevisiae. Biochim Biophys Acta 930: 220–229PubMedCrossRefGoogle Scholar
  26. 26.
    Kim SC, Sprung R, Chen Y, Xu Y, Ball H, Pei J, Cheng T, Kho Y, Xiao H, Xiao L, Grishin NV, White M, Yang XJ, Zhao Y (2006) Substrate and functional diversity of lysine acetylation revealed by a proteomics survey. Mol Cell 23:607–618PubMedCrossRefGoogle Scholar
  27. 27.
    Choudhary C, Kumar C, Gnad F, Nielsen ML, Rehman M, Walther TC, Olsen JV, Mann M (2009) Lysine acetylation targets protein complexes and co-regulates major cellular functions. Science 325(5942):834–840PubMedCrossRefGoogle Scholar
  28. 28.
    Zhao S, Xu W, Jiang W, Yu W, Lin Y, Zhang T, Yao J, Zhou L, Zeng Y, Li H, Li Y, Shi J, An W, Hancock SM, He F, Qin L, Chin J, Yang P, Chen X, Lei Q, Xiong Y, Guan KL (2010) Regulation of cellular metabolism by protein lysine acetylation. Science 327(5968): 1000–1004PubMedCrossRefGoogle Scholar
  29. 29.
    Vijg J, Campisi J (2008) Puzzles, promises and a cure for ageing. Nature 454(7208): 1065–1071PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2013

Authors and Affiliations

  • Jin-ying Lu
    • 1
  • Yu-yi Lin
    • 2
  • Jef D. Boeke
    • 3
  • Heng Zhu
    • 4
    • 5
  1. 1.Department of Laboratory MedicineNational Taiwan University Hospital, Institute of Molecular Medicine, College of Medicine, National Taiwan UniversityTaipeiTaiwan
  2. 2.Department of OncologyNational Taiwan University Hospital, Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan UniversityTaipeiTaiwan
  3. 3.Department of Molecular Biology and Genetics, The High Throughput Biology CenterJohn Hopkins University School of MedicineBaltimoreUSA
  4. 4.Departments of Pharmacology, The High Throughput Biology CenterJohn Hopkins University School of MedicineBaltimoreUSA
  5. 5.Department Molecular Sciences, The High Throughput Biology CenterJohn Hopkins University School of MedicineBaltimoreUSA

Personalised recommendations