The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3

Anne M. Cieniewicz, Linley Moreland, Alison E. Ringel, Samuel G. Mackintosh, Ana Raman, Tonya M. Gilbert, Cynthia Wolberger, Alan J. Tackett, Sean Dixon Taverna

Research output: Contribution to journalArticle

Abstract

In yeast, the conserved histone acetyltransferase (HAT) Gcn5 associates with Ada2 and Ada3 to form the catalytic module of the ADA and SAGA transcriptional coactivator complexes. Gcn5 also contains an acetyl-lysine binding bromodomain that has been implicated in regulating nucleosomal acetylation in vitro, as well as at gene promoters in cells. However, the contribution of the Gcn5 bromodomain in regulating site specificity of HAT activity remains unclear. Here, we used a combined acid-urea gel and quantitative mass spectrometry approach to compare the HAT activity of wild-type and Gcn5 bromodomain- mutant ADA subcomplexes (Gcn5-Ada2-Ada3). Wild-type ADA subcomplex acetylated H3 lysines with the following specificity; H3K14 > H3K23 > H3K9 ≈ H3K18 > H3K27 > H3K36. However, when the Gcn5 bromodomain was defective in acetyl-lysine binding, the ADA subcomplex demonstrated altered site-specific acetylation on free and nucleosomal H3, with H3K18ac being the most severely diminished. H3K18ac was also severely diminished on H3K14R, but not H3K23R, substrates in wildtype HAT reactions, further suggesting that Gcn5-catalyzed acetylation of H3K14 and bromodomain binding to H3K14ac are important steps preceding H3K18ac. In sum, this work details a previously uncharacterized cross-talk between the Gcn5 bromodomain "reader" function and enzymatic HAT activity that might ultimately affect gene expression. Future studies of how mutations in bromodomains or other histone post-translational modification readers can affect chromatinlated enzymatic activities will yield unprecedented insight into a potential "histone/epigenetic code." MS data are available via ProteomeXchange with identifier PXD001167.

Original languageEnglish (US)
Pages (from-to)2896-2910
Number of pages15
JournalMolecular and Cellular Proteomics
Volume13
Issue number11
DOIs
StatePublished - Nov 1 2014

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Histone Acetyltransferases
Acetylation
Histones
Lysine
Histone Code
Post Translational Protein Processing
Epigenomics
Gene expression
Yeast
Mass spectrometry
Urea
Mass Spectrometry
Genes
Yeasts
Gels
Gene Expression
Mutation
Acids
Substrates

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Analytical Chemistry

Cite this

Cieniewicz, A. M., Moreland, L., Ringel, A. E., Mackintosh, S. G., Raman, A., Gilbert, T. M., ... Taverna, S. D. (2014). The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3. Molecular and Cellular Proteomics, 13(11), 2896-2910. https://doi.org/10.1074/mcp.M114.038174

The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3. / Cieniewicz, Anne M.; Moreland, Linley; Ringel, Alison E.; Mackintosh, Samuel G.; Raman, Ana; Gilbert, Tonya M.; Wolberger, Cynthia; Tackett, Alan J.; Taverna, Sean Dixon.

In: Molecular and Cellular Proteomics, Vol. 13, No. 11, 01.11.2014, p. 2896-2910.

Research output: Contribution to journalArticle

Cieniewicz, AM, Moreland, L, Ringel, AE, Mackintosh, SG, Raman, A, Gilbert, TM, Wolberger, C, Tackett, AJ & Taverna, SD 2014, 'The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3', Molecular and Cellular Proteomics, vol. 13, no. 11, pp. 2896-2910. https://doi.org/10.1074/mcp.M114.038174
Cieniewicz AM, Moreland L, Ringel AE, Mackintosh SG, Raman A, Gilbert TM et al. The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3. Molecular and Cellular Proteomics. 2014 Nov 1;13(11):2896-2910. https://doi.org/10.1074/mcp.M114.038174
Cieniewicz, Anne M. ; Moreland, Linley ; Ringel, Alison E. ; Mackintosh, Samuel G. ; Raman, Ana ; Gilbert, Tonya M. ; Wolberger, Cynthia ; Tackett, Alan J. ; Taverna, Sean Dixon. / The bromodomain of gcn5 regulates site specificity of lysine acetylation on histone H3. In: Molecular and Cellular Proteomics. 2014 ; Vol. 13, No. 11. pp. 2896-2910.
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AU - Raman, Ana

AU - Gilbert, Tonya M.

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AU - Tackett, Alan J.

AU - Taverna, Sean Dixon

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