The Bromodomain of Gcn5 Regulates Site Specificity of Lysine Acetylation on Histone H3

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

doi:10.1074/mcp.m114.038174

Cited by 66 publications

(64 citation statements)

References 93 publications

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“…To examine whether recognition of the H3K4me3 modification changes the intrinsic lysine specificity of the Gcn5 subunit, which preferentially catalyzes H3K14 acetylation in vitro (29,39) and H3K9 acetylation in vivo (20,40), we probed the reaction products using antibodies that recognize specific acetylated lysine residues. Antibodies to the H3K9ac and H3K14ac modifications recognize five bands on acid-urea gels, corresponding to one through five modification sites per histone, whereas antibodies to H3K18ac and H3K23ac recognize only the top four bands (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Nucleosome competition reveals processive acetylation by the SAGA HAT module

Ringel

Cieniewicz

Taverna

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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The Spt-Ada-Gcn5 acetyltransferase (SAGA) coactivator complex hyperacetylates histone tails in vivo in a manner that depends upon histone 3 lysine 4 trimethylation (H3K4me3), a histone mark enriched at promoters of actively transcribed genes. SAGA contains a separable subcomplex known as the histone acetyltransferase (HAT) module that contains the HAT, Gcn5, bound to Sgf29, Ada2, and Ada3. Sgf29 contains a tandem Tudor domain that recognizes H3K4me3-containing peptides and is required for histone hyperacetylation in vivo. However, the mechanism by which H3K4me3 recognition leads to lysine hyperacetylation is unknown, as in vitro studies show no effect of the H3K4me3 modification on histone peptide acetylation by Gcn5. To determine how H3K4me3 binding by Sgf29 leads to histone hyperacetylation by Gcn5, we used differential fluorescent labeling of histones to monitor acetylation of individual subpopulations of methylated and unmodified nucleosomes in a mixture. We find that the SAGA HAT module preferentially acetylates H3K4me3 nucleosomes in a mixture containing excess unmodified nucleosomes and that this effect requires the Tudor domain of Sgf29. The H3K4me3 mark promotes processive, multisite acetylation of histone H3 by Gcn5 that can account for the different acetylation patterns established by SAGA at promoters versus coding regions. Our results establish a model for Sgf29 function at gene promoters and define a mechanism governing crosstalk between histone modifications.T he different patterns of histone posttranslational modifications distributed across the genome are proposed to constitute a "histone code" that orchestrates distinct transcriptional programs by recruiting specific effector proteins (1-4). The phenomenon of histone code "crosstalk," whereby one type of histone modification directs the establishment of another, or through which several histone modifications are recognized in tandem, has emerged as an important and widespread mechanism regulating chromatin-templated processes (5, 6). The multifunctional complexes that activate transcription are thought to mediate crosstalk through "reader" domains that recognize particular chromatin marks as well as through catalytic subunits that deposit or remove histone modifications (5,7,8). As a result, distinct combinations of histone posttranslational modifications that correlate with transcriptional output cluster across the genome (9-11). High levels of histone 3 lysine 4 trimethylation (H3K4me3) and histone H3 hyperacetylation are present at the promoters of actively transcribed genes (9,11,12), and multiple lines of evidence support the existence of regulatory mechanisms coupling the two modifications. Studies using tandem mass spectrometry to sequence whole histone tails have shown that H3K4me3 is highly correlated with hyperacetylation of the same H3 tail (13,14). Deleting the yeast Set1 methyltransferase, which trimethylates H3K4, leads to dramatically lower levels of histone H3 tail acetylation overall (13, 15). These results are consistent with ...

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Section: Resultsmentioning

confidence: 99%

“…6). In the absence of H3K4 trimethylation, the HAT module likely acetylates histones according to the intrinsic lysine specificity of Gcn5, which vastly favors modification at the H3K14 position (29,39,51). At promoters, however, recognition of the H3K4me3 mark by Sgf29 promotes processive, multisite acetylation by the HAT module.…”

Section: Discussionmentioning

confidence: 99%

Nucleosome competition reveals processive acetylation by the SAGA HAT module

Ringel

Cieniewicz

Taverna

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…A study combining in vitro acid-urea gel and quantitative mass spectrometry approach to measure the activity of yeast Gcn5 incorporated in the partial HAT module of the ySAGA and Ada complexes containing Gcn5-Ada2-Ada3 showed that Gcn5 acetylated free histone H3 with the following efficiency: H3K14 Ͼ H3K23 Ͼ H3K9 Ϸ H3K18 Ͼ H3K27 Ͼ H3K36 (23). Purified yeast and human SAGA complexes were shown in vitro to acetylate H3 mainly at position Lys-14, but also Lys-9, Lys-18, and Lys-23 to some extent when using different preacetylated H3 tail peptides (24,25).…”

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confidence: 99%

Subunits of ADA-two-A-containing (ATAC) or Spt-Ada-Gcn5-acetyltrasferase (SAGA) Coactivator Complexes Enhance the Acetyltransferase Activity of GCN5

Riss

Scheer

Joint

et al. 2015

Journal of Biological Chemistry

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Background: Histone acetyltransferases (HATs) incorporated in large multiprotein complexes are involved in a wide variety of cellular processes, including transcription regulation. Results: Subunits of HAT complexes enhance the enzymatic activity of their catalytic subunits. Conclusion: The activity, but not the specificity of HAT complexes, is stimulated by the corresponding subunits. Significance: We gained important insights into histone acetylation function and specificity of HAT complexes.

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“…This is not a comprehensive list of histone PTMs, but rather represents the most abundant bulk level histone PTMs that can be identified with high confidence. Further separation of histones based on the PTM state (as we have done previously (31,33)) can lead to more comprehensive lists of PTMs, which will be the focus of future work. Histone PTMs having a p value less than 0.1 were considered of interest and included H3K27me3 and H4K20me (supplemental Tables 7-9).…”

Section: Histone H3k27me3 Is Elevated In Late Stage Melanomamentioning

confidence: 99%

“…In this study, we utilized a label-free precursor ion intensity approach for a bottom-up analysis of histone PTMs (30). To increase sequence coverage for the lysine-and arginine-rich histones that yield very small peptides upon trypsin digestion, we used d 6 -acetic anhydride to chemically label unmodified and monomethylated lysines with an isotopically heavy acetyl group, which blocks trypsin activity and thereby increases sequence coverage following trypsin digestion (31,32). By increasing the sequence coverage, we can both increase detection of potential sites of modification as well as obtain larger peptides containing multiple sites of potential modification thereby gaining insight into combinatorial histone modifications on individual histone molecules.…”

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confidence: 99%

Quantitative Histone Mass Spectrometry Identifies Elevated Histone H3 Lysine 27 (Lys27) Trimethylation in Melanoma

Sengupta

Byrum

Avaritt

et al. 2016

Molecular & Cellular Proteomics

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Normal cell growth is characterized by a regulated epigenetic program that drives cellular activities such as gene transcription, DNA replication, and DNA damage repair. Perturbation of this epigenetic program can lead to events such as mis-regulation of gene transcription and diseases such as cancer. To begin to understand the epigenetic program correlated to the development of melanoma, we performed a novel quantitative mass spectrometric analysis of histone post-translational modifications mis-regulated in melanoma cell culture as well as patient tumors. Aggressive melanoma cell lines as well as metastatic melanoma were found to have elevated histone H3 Lys 27 trimethylation (H3K27me3) accompanied by overexpressed methyltransferase EZH2 that adds the specific modification. The altered epigenetic program that led to elevated H3K27me3 in melanoma cell culture was found to directly silence transcription of the tumor suppressor genes RUNX3 and E-cadherin. The EZH2-mediated silencing of RUNX3 and E-cadherin transcription was also validated in advanced stage human melanoma tissues. This is the first study focusing on the detailed epigenetic mechanisms leading to EZH2-mediated silencing of RUNX3 and E-cadherin tumor suppressors in melanoma. This study underscores the utility of using high resolution mass spectrometry to identify misregulated epigenetic programs in diseases such as cancer, which could ultimately lead to the identification of biological markers for diagnostic and prognostic applications. Melanoma is a deadly variety of skin cancer, accounting for 75% of skin cancer-related deaths. In 2015, melanoma is expected to be the fifth most common cancer in men and the seventh most common cancer in women. According to the World Health Organization, it is estimated that melanoma will result in the death of around 65,000 people globally and 9940 people in the United States in 2015. The high mortality rate associated with metastatic melanoma suggests a lack of efficient diagnostic and prognostic biomarkers (1). EZH2 1 expression has often been positively correlated to the progression of different types of cancer (2, 3). Increased expression of EZH2 has been identified in melanoma tissues (4) as well as prostate, breast, bladder, and liver cancers and has been recognized as a prognostic marker for aggressive prostate and breast cancer (5, 6). EZH2 is a histone modifier that functions as the catalytic component of the Polycomb Repressive Complex 2 (PRC2) (7,8). It is a lysine methyltransferase and promotes the addition of the repressive marker histone H3K27me2/me3 to target chromatin, thereby inducing chromatin compaction and transcriptional repression. Chromatin condensation/compaction leads to transcriptional repression by restricting access to transcriptional regulators like RNA polymerase II and other transcription-associated factors. Hence, silencing of tumor suppressor genes by H3K27me3 is implicated in the initiation and advancement of different types of cancer (9 -11). H3K27me3-silenced tumor suppressor gene...

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The Bromodomain of Gcn5 Regulates Site Specificity of Lysine Acetylation on Histone H3

Cited by 66 publications

References 93 publications

Nucleosome competition reveals processive acetylation by the SAGA HAT module

Nucleosome competition reveals processive acetylation by the SAGA HAT module

Subunits of ADA-two-A-containing (ATAC) or Spt-Ada-Gcn5-acetyltrasferase (SAGA) Coactivator Complexes Enhance the Acetyltransferase Activity of GCN5

Quantitative Histone Mass Spectrometry Identifies Elevated Histone H3 Lysine 27 (Lys27) Trimethylation in Melanoma

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