Histone acetyltransferase activity of yeast Gcn5p is required for the activation of target genes in vivo

Abstract

Gcn5p is a transcriptional coactivator required for correct expression of various genes in yeast. Several transcriptional regulators, including Gcn5p, possess intrinsic histone acetyltransferase (HAT) activity in vitro. However, whether the HAT activity of any of these proteins is required for gene activation remains unclear. Here, we demonstrate that the HAT activity of Gcn5p is critical for transcriptional activation of target genes in vivo. Core histones are hyperacetylated in cells overproducing functional Gcn5p, and promoters of Gcn5p-regulated genes are associated with hyperacetylated histones upon activation by low-copy Gcn5p. Point mutations within the Gcn5p catalytic domain abolish both promoter-directed histone acetylation and Gcn5p-mediated transcriptional activation. These data provide the first in vivo evidence that promoter-specific histone acetylation, catalyzed by functional Gcn5p, plays a critical role in gene activation.

Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes

“…Histone acetylation is perhaps the best studied modification and has long been associated with gene activation. The molecular basis for this association became apparent when it was discovered that the transcriptional coactivator Gcn5 possessed histone acetyltransferase activity and that transcriptional activity of Gcn5-dependent genes in a yeast cell correlated tightly with Gcn5's HAT activity [2][3][4].…”

Chromatin remodeling and transcriptional activation: the cast (in order of appearance)

“…While its elementary particle, the nucleosome (Kornberg and Thomas, 1974), is ubiquitous (Noll, 1974), chromatin over a given DNA locus can assume a great variety of markedly distinct structural states in vivo (Hebbes et al, 1994(Hebbes et al, , 1988Tumbar et al, 1999;Wu, 1980;Zaret and Yamamoto, 1984): after all, many di erent buildings can be created using the same bricks. There is very strong correlative evidence connecting chromatin structure of a speci®c locus and the level of its transcriptional activity (for example, in addition to the studies just cited, Bone et al, 1994;Braunstein et al, 1993;Jeppesen and Turner, 1993;Kuo et al, 1998). In comforting parallel, for certain loci in some model systems there is equally strong biochemical and genetic evidence that implicate speci®c protein complexes both in e ecting chemical or structural transitions in chromatin and regulating transcription over target genes (for example, Goldmark et al, 2000;Gregory et al, 1999;Moreira and Holmberg, 1999;Rundlett et al, 1998;Zhang et al, 1998).…”

“…This celebrity is due to the strong positive correlation between levels of acetylation of speci®c loci in the genome and their transcriptional activity (Braunstein et al, 1993;Hebbes et al, 1988;Jeppesen and Turner, 1993;Kuo et al, 1998;Parekh and Maniatis, 1999), and is also a result of the nearomnipresence of histone acetyltransferases (HATs) (Brownell et al, 1996;Sterner and Berger, 2000) and deacetylases (HDACs) (Ng and Bird, 2000;Taunton et al, 1996) in transcriptional control pathways.…”

“…13 acetylated lysines per octamer, i.e., on just under 50% of all possible acetylation sites in each nucleosome! Subsequent genetic analysis (Kuo et al, 2000;Vogelauer et al, 2000) implicated the HATs Gcn5p and Esa1p in this`genome-wide' acetylation, and indicated that promoter-speci®c increases in core histone acetylation that occur concomitantly with gene activation (Krebs et al, 1999;Kuo et al, 1998) are the result of targeting these HATs by promoter-bound regulators to increase the localized concentration of HAT enzymes at gene promoters.`Nontargeted' action by yeast HATs illuminates earlier data that HDAC inhibitors such as trichostatin A (TSA) can upregulate basal activity of speci®c promoters that do not appear to have HATs targeted to them (Urnov et al, 2000). Thus, the current model is that budding yeast chromatin is under constant attack from untargeted HATs and HDACs, and these create a high steadystate level of tail acetylation ( Figure 1).…”

“…It should not be inferred, however, that targeted acetylation is not required for gene activation in yeast: such HATs as Gcn5p hyperacetylate chromatin in vivo over target promoters (Krebs et al, 1999;Kuo et al, 1998Kuo et al, , 2000, and the lysines in the core histone tails are bona ®de substrates for such action (Zhang et al, 1998) ± this latter study was a technical tour de force, because it presented evidence that`genetic hyperacetylation of histones' (i.e., the mutation of Gcn5-targeted lysines to a non-charged glutamine) obviates the requirement for the Gcn5p HAT in activation of a speci®c gene! In general, Gcn5p is required for the proper upregulation of a signi®cant number of yeast genes (Lee et al, 2000), and its HAT domain is essential for this phenomenon (Kuo et al, 1998;Wang et al, 1998).…”