Nucleosome competition reveals processive acetylation by the SAGA HAT module

Abstract

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 ...

Distinct requirements of linker DNA and transcriptional activators in promoting SAGA-mediated nucleosome acetylation

“…It should be noted that release and preferential rebinding of acetylated nucleosomes might mimic nucleosome acetylation without release. However, a published study is consistent with committed binding between multiple nucleosome acetylations (47), and thus we interpret the burst-phase kinetics . Effect of activator protein on SAGA activity.…”

Structural basis for COMPASS recognition of an H2B-ubiquitinated nucleosome

“…H2B-Ub and H3K4 di-and tri-methylation are strongly associated 15 with active transcription in both yeast and humans (Barski et al, 2007;Jung et al, 2012;16 Liu et al, 2005;Minsky et al, 2008;Pokholok et al, 2005;Santos-Rosa et al, 2002;17 Shieh et al, 2011;Steger et al, 2008). H3K4 methylation can serve as a recruitment 18 signal for various transcription activators (Sims et al, 2007;Vermeulen et al, 2010;19 Vermeulen et al, 2007;Wysocka et al, 2006) including SAGA, whose acetyltransferase 20 activity is stimulated by H3K4 methylation (Bian et al, 2011;Ringel et al, 2015). 21…”

KATapulting toward Pluripotency and Cancer

“…The SAGA complex contains four functional units that elegantly control transcriptional activation, telomere maintenance 72 , mRNA export 73 and DNA repair 74 . One module consists of the acetyltransferase unit (Gcn5, Ada2b, Ada3, Sgf29) that harbors KAT activity and facilitates SAGA recruitment to H3K4me2/3 sites via the tandem Tudor domains of Sgf29 75; 76 and/or to acetyl lysine residues through the bromodomain of Gcn5 77 . A second enzymatic module, called the deubiquitination (DUB) module (Usp22, Atxn7, Atxn7L3 and Eny2) promotes H2BK120 deubiquitination during transcription 78 .…”