Two Different Drosophila ADA2 Homologues Are Present in Distinct GCN5 Histone Acetyltransferase-Containing Complexes

Abstract

We have isolated a novel Drosophila (d) gene coding for two distinct proteins via alternative splicing: a homologue of the yeast adaptor protein ADA2, dADA2a, and a subunit of RNA polymerase II (Pol II), dRPB4. Moreover, we have identified another gene in the Drosophila genome encoding a second ADA2 homologue (dADA2b). The two dADA2 homologues, as well as many putative ADA2 homologues from different species, all contain, in addition to the ZZ and SANT domains, several evolutionarily conserved domains. The dada2a/rpb4 and dada2b genes are differentially expressed at various stages of Drosophila development. Both dADA2a and dADA2b interacted with the GCN5 histone acetyltransferase (HAT) in a yeast two-hybrid assay, and dADA2b, but not dADA2a, also interacted with Drosophila ADA3. Both dADA2s further potentiate transcriptional activation in insect and mammalian cells. Antibodies raised either against dADA2a or dADA2b both immunoprecipitated GCN5 as well as several Drosophila TATA binding protein-associated factors (TAFs). Moreover, following glycerol gradient sedimentation or chromatographic purification combined with gel filtration of Drosophila nuclear extracts, dADA2a and dGCN5 were detected in fractions with an apparent molecular mass of about 0.8 MDa whereas dADA2b was found in fractions corresponding to masses of at least 2 MDa, together with GCN5 and several Drosophila TAFs. Furthermore, in vivo the two dADA2 proteins showed different localizations on polytene X chromosomes. These results, taken together, suggest that the two Drosophila ADA2 homologues are present in distinct GCN5-containing HAT complexes.Transcription in eukaryotes is a tightly regulated, multistep process. General transcription factors, gene specific transcriptional activators, and several different cofactors are necessary to access specific loci in the context of eukaryotic chromatin to allow precise initiation of RNA polymerase II (Pol II) transcription. One of the most appealing questions in eukaryotic transcription is how activators transmit their signals to the general transcription machinery to stimulate transcription.Posttranslational modifications of nucleosomal histones have been correlated with the function of chromatin in transcription activation or repression (18, 34). One of the most extensively studied modifications is the acetylation of the highly conserved amino-terminal histone tails. The steady-state level of acetylation of histone proteins is accomplished by the action of histone acetyltransferases (HATs) and histone deacetylases (9, 37). Acetylation affects higher-order folding of chromatin fibers and histone-nonhistone protein interactions (31, 32). Thus, it can increase the affinity of transcription factors for nucleosomal DNA (40,61).A large number of recent studies have provided a direct molecular link between histone acetylation and transcriptional activation (reviewed in references 9 and 30). In these reports, it has been shown that several previously identified coactivators and adaptors of transcription possess i...

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

“…Interestingly, the yAda2 protein has two paralogues in metazoans: ADA2a and ADA2b (28,29). Although ADA2b is part of the HAT module of the SAGA complex, ADA2a was shown to be a subunit of the HAT module of the ATAC complex together with GCN5, ADA3, and SGF29 (18,28,29).…”

The role of human ADA2a in the regulation of p53 acetylation and stability

“…No role for ADA2 independent of Gcn5 has been characterized in Saccharomyces cerevisiae, although multiple forms of ADA2 exist in Drosophila melanogaster [16,22], plants [23], mice, and humans [17]. In Drosophila, ADA2a and ADA2b are required for normal development but have distinct functions [24].…”

“…Recently, several groups have reported that the Drosophila genome contains 2 distinct genes encoding ADA2 homologs: ADA2a and ADA2b. Biochemical characterization of the 2 ADA2 proteins demonstrated that both interact with the HAT/ GCN5 complex and participate in transcriptional activation [16]. However, functional differences between the 2 human ADA2 proteins, hADA2a and hADA2b, have been recently reported [17].…”

TAF10 and TAF10b partially redundant roles duringDrosophila melanogastermorphogenesis

“…It has already been demonstrated that dTAF10 is part of the dSAGA complex, while dTAF10b is a component of the dTFIID complex. 16,17,20 We also checked available Drosophila protein-protein interaction databases and collected the interaction partners of dTAF10 and dTAF10b. The results were visualized by using Cytoscape software.…”

“…The two dTAF10 proteins show different expression patterns in neuronal tissues during embryonic development and are present in different multiprotein complexes -dSAGA and dTFIID, respectively. [16][17][18] Recently, it has been reported that dTAF10 and dTAF10b have synergistic roles in the regulation of Drosophila morphogenesis and they are required for the activation of Halloween genes involved in ecdysone biosynthesis. [18][19][20] Taken together, these observations support the idea that functionally different TAFcontaining complexes participate in gene regulation.…”