In cells, DNA is complexed with nucleosomes to create the higher-order repressive chromatin structure characteristic of chromosomes. Nucleosomal associations with DNA are dynamic, with modifications of histone components occurring during DNA repair and recombination, and regulation of transcription. In particular, hyperacetylation of histone tails is correlated with transcriptionally active chromatin, and hypoacetylated histones are found within silent heterochromatic DNA. Transcriptional adaptors, or ADAs, are evolutionarily conserved proteins that are required for full function of certain transcriptional activators in vivo. One of these adaptors, GCN5, is newly recognized as a nuclear histone acetyltransferase, suggesting that adaptors function in part to remodel chromatin during transcriptional activation. Recombinant GCN5 acetylates only free histones, however, as a component of two high molecular weight native yeast complexes, it acquires the ability to acetylate histones incorporated into nucleosomes. The yeast complexes contain additional factors required for transcriptional activation. The proposed research will characterize the composition of these complexes, to study their structure and functions, and to determine their physiological relevance. The hypothesis of the work is that multiple regulatory functions that potentiate transcriptional activation are integrated in these complexes. The goal of the research is to determine how these functions operate and are integrated to activate transcription. The function of the yeast nucleosomal acetylation complexes will be analyzed in transcriptional activation and nucleosome remodeling in vivo and in vitro. The structure and composition of the complexes will be studied. Finally the ability of the complexes to physically associate with other key transcription factors will be tested. The yeast complexes containing GCN5 and possessing nucleosomal acetylation function may have a key regulatory role in gene expression. GCN5 is conserved through evolution and additional transcriptional regulatory factors have been identified as histone acetyltransferases. Thus, deciphering the mechanism of action of GCN5 in transcriptional activation will contribute to an increased understanding of how regulation goes awry in many diseases states, including cancer.
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