DrosophilaAda2b Is Required for Viability and Normal Histone H3 Acetylation

Abstract

Regulation of chromatin through histone acetylation is an important step in gene expression. The Gcn5 histone acetyltransferase is part of protein complexes, e.g., the SAGA complex, that interact with transcriptional activators, targeting the enzyme to specific promoters and assisting in recruitment of the basal RNA polymerase transcription machinery. The Ada2 protein directly binds to Gcn5 and stimulates its catalytic activity. Drosophila contains two Ada2 proteins, Drosophila Ada2a (dAda2a) and dAda2b. We have generated flies that lack dAda2b, which is part of a Drosophila SAGA-like complex. dAda2b is required for viability in Drosophila, and its deletion causes a reduction in histone H3 acetylation. A global hypoacetylation of chromatin was detected on polytene chromosomes in dAda2b mutants. This indicates that the dGcn5-dAda2b complex could have functions in addition to assisting in transcriptional activation through gene-specific acetylation. Although the Drosophila p53 protein was previously shown to interact with the SAGA-like complex in vitro, we find that p53 induction of reaper gene expression occurs normally in dAda2b mutants. Moreover, dAda2b mutant animals show excessive p53-dependent apoptosis in response to gamma radiation. Based on this result, we speculate that dAda2b may be necessary for efficient DNA repair or generation of a DNA damage signal. This could be an evolutionarily conserved function, since a yeast ada2 mutant is also sensitive to a genotoxic agent.The Ada2 protein was first described for yeast, in which its inactivation could relieve the toxicity caused by overexpression of the strong transcriptional activation domain in the viral VP16 protein (6). This provided one of the earliest pieces of evidence for the existence of transcriptional adapters or coactivators. Ada2 proteins were subsequently shown to exist in protein complexes containing the Gcn5 histone acetyltransferase (23). Histone acetylation has been linked to transcriptional activation for a long time, given the observation that actively transcribed regions tend to be hyperacetylated, whereas transcriptionally silent regions generally are hypoacetylated (3). With the identification of the first histone acetyltransferase (HAT) as the Tetrahymena homolog of Gcn5 (13) and the subsequent demonstration that other transcriptional regulators, such as the coactivators p300 and CBP, steroid receptor coactivators, and the TFIID subunit TAF1, also possess intrinsic HAT activity, the link between acetylation and transcription was firmly established (12). In addition, an important role for histone acetylation in DNA replication and repair is emerging (27,38).Gcn5 is evolutionarily conserved and is present in several multiprotein complexes, such as the SAGA and ADA complexes in yeast and the PCAF, STAGA, and TFTC complexes in mammals (17). In yeast, Gcn5 is contained in a trimeric subcomplex with the Ada2 and Ada3 proteins within SAGA and ADA complexes (21,24). Ada2 binds to Gcn5, and deletion of Ada2 results in a loss of Gcn5 from bot...

Distinct GCN5/PCAF-containing complexes function as co-activators and are involved in transcription factor and global histone acetylation

“…Similar to GCN5, both Drosophila Ada2 genes are essential. Ada2a homozygote mutants are late-larva lethal, while Ada2b mutant animals die later at the end of the prepupal phase (Qi et al, 2004;Pankotai et al, 2005). Mutations of the Ada2b gene result in a loss of acetylation of residues K9 and K14 on H3, while Ada2a mutations only affect acetylation of K5 and K12 residues on histone H4 when tested on polytene chromosomes (Qi et al, 2004;Pankotai et al, 2005;Ciurciu et al, 2006).…”

Nipped-A, the Tra1/TRRAP Subunit of the Drosophila SAGA and Tip60 Complexes, Has Multiple Roles in Notch Signaling during Wing Development

“…All three mutant alleles were generated by P element excision and were compared to the original viable P element-bearing chromosome [EP(3)3412] used to generate them (44; T Kusch, personal communication). The Ada2b 1 and Ada2b 2 alleles have large deletions (1.1 and 2.8 kbp) that remove the transcription start site and much of the gene, and homozygotes die during pupariation (44). The Ada2b HD-5 allele, an unpublished allele provided by Thomas Kusch and Jerry Workman (Stowers Institute), appears to be hypomorphic because a small percentage of homozygotes eclose but die shortly thereafter.…”

A dSAGA specifikus hiszton acetiláció génműködés szabályozásban betöltött funkciójának vizsgálata

“…Az ADA2 fehérjék sokszínű funkciójára utal, hogy a Drosophila Ada2b génről több mRNS és fehérje izoforma termelődik [73]. Kutatócsoportunk korábban már igazolta, hogy az egyedfejlődés főbb szakaszaiban (embrió, lárva, báb, imágó állapot) a dAda2b génről alternatív splicing során valójában két mRNS forma (dAda2bS és dAda2bL) képződik [72].…”

“…) mutáció is a báb fejlődésének korai szakaszában okoz letalitást, így feltételezhetjük, hogy a dADA2b fehérjék fontos funkciót töltenek be a metamorfózis folyamatában [72] [73].…”