Expanded Lysine Acetylation Specificity of Gcn5 in Native Complexes

Abstract

The coactivator/adaptor protein Gcn5 is a conserved histone acetyltransferase, which functions as the catalytic subunit in multiple yeast transcriptional regulatory complexes. The ability of Gcn5 to acetylate nucleosomal histones is significantly reduced relative to its activity on free histones, where it predominantly modifies histone H3 at lysine 14. However, the association of Gcn5 in multisubunit complexes potentiates its nucleosomal histone acetyltransferase activity. Here, we show that the association of Gcn5 with other proteins in two native yeast complexes, Ada and SAGA (Spt-Ada-Gcn5-acetyltransferase), directly confers upon Gcn5 the ability to acetylate an expanded set of lysines on H3. Furthermore Ada and SAGA have overlapping, yet distinct, patterns of acetylation, suggesting that the association of specific subunits determines site specificity.In the eukaryotic nucleus DNA is packaged by histones into nucleosomes, the repeating subunits of chromatin. The nucleosome core particle contains about 146 base pairs of DNA and two of each of the histones H2A, H2B, H3, and H4. Packaging of DNA into chromatin suppresses transcription, at least in part by occluding the binding of transcriptional activators and basal transcription machinery to their cognate DNA sites (1, 2) and by blocking transcriptional elongation (3). A number of chromatin remodeling activities have been isolated that assist activators to bind to their sites by promoting a localized alteration in chromatin structure (4).In addition to these activities, post-translational modifications of the core histones within the nucleosome have also been linked to the transcriptional capacity of chromatin (5, 6). The acetylation of the highly conserved lysine residues within the amino-terminal domains of all four core histones has been extensively studied (7). Histone acetylation has been associated with the interaction of non-histone proteins with histones (8, 9), histone deposition and nucleosome assembly (10), higher order packing of chromatin (11,12), and the transcriptional activity of cellular chromatin (5, 13). Correlations between transcription and acetylation are reinforced by studies demonstrating that at least some active chromosomal domains are hyperacetylated (14), while inactive or heterochromatin domains are associated with hypoacetylated histones (15-18). However these general correlations do not hold true when specific acetylated H4 isoforms are monitored indicating that particular patterns of lysine acetylation rather than bulk acetylation is of functional importance (15,19,20). Therefore, an important goal in studying this phenomenon has been the isolation of native activities that generate such specific patterns of acetylation.The steady state level of acetylation of histones is a balance between the action of histone acetyltransferases (HATs) 1 and histone deacetylases. These activities are often found to be associated with large multisubunit protein complexes and contain known regulators of transcription (21). A number of previously ...

Developmental potential of Gcn5−/− embryonic stem cells in vivo and in vitro

“…Gcn5 acetylates histones H3 and H2B (Grant et al, 1997(Grant et al, , 1999. H3 acetylation is decreased approximately twofold in undifferentiated Gcn5 Ϫ/Ϫ cells (data not shown), indicating that Gcn5 is a major H3 HAT activity in these cells, but it is not the only one.…”

A New Coactivator Function for Zac1's C2H2 Zinc Finger DNA-Binding Domain in Selectively Controlling PCAF Activity

“…In support of this view, Zac1 is here shown to increase the rate of acetylation of histone H4 versus H3 due to changes in PCAF's substrate affinities and catalytic activities. In this context, it is important to note that previous studies showed that association with other factors leads to a context-dependent acetylation pattern for the PCAF-related acetyltransferase Gcn5 (14). Among the factors that associate with Gcn5 are the adaptor proteins Ada2, which potentiates Gcn5 catalytic activity, and Ada3, which facilitates nucleosomal acetylation and an expanded lysine specificity (2).…”

Catalytic activation of histone acetyltransferase Rtt109 by a histone chaperone

“…HATs are almost exclusively found within larger multisubunit complexes with accessory proteins that modulate enzymatic activity and direct substrate specificity (2)(3)(4). For example, the yeast HATs Esa1 (KAT5) and Gcn5 (KAT2) alone are ineffective catalysts toward nucleosomal histones; both enzymes require association with other protein subunits for efficient acetyl transfer on nucleosomal substrates (5)(6)(7). Despite the prevalent reports of multisubunit HAT complexes, the molecular mechanisms by which accessory proteins regulate the acetyltransferases are largely unknown.…”