Antagonistic Gcn5-Hda1 interactions revealed by mutations to the Anaphase Promoting Complex in yeast

Using Northerns, Westerns, reverse transcriptase PCR (rtPCR), chromatin immunoprecipitation (ChIP), and mutant phenotype suppression analysis, we observed that Hda1 and Gcn5 appear to compete for recruitment to promoters. We observed that the presence of Hda1 can partially occlude the binding of Gcn5 to the same promoter. Occlusion of Gcn5 recruitment to these promoters involved Hda1 and Tup1. Using sequential ChIP we show that Hda1 and Tup1 likely form complexes at these promoters, and that complex formation can be increased by deleting GCN5.Our data suggests large Gcn5 and Hda1 containing complexes may compete for space on promoters that utilize the Ssn6/Tup1 repressor complex. We predict that in apc5CA cells the accumulation of an APC target may compensate for the loss of both GCN5 and HDA1.Eukaryotic genetic information is packaged into chromatin, a highly organized and dynamic protein-DNA complex. The fundamental unit of chromatin, the nucleosome, is an octameric structure composed of two copies of each of the four core histones (an H3/H4 tetramer and two H2A/H2B dimers), surrounded by approximately 146 bp of DNA [1,2]. Many cellular processes depend on modifications of both DNA and histones within nucleosomes [3,4]. Modification of chromatin by histone acetyltransferases (HATs) and histone deacetylases (HDACs) play key roles in transcriptional regulation [5-9]. Post-translational acetylation of the highly conserved lysines within the N-terminal tail domains of the core histones is strongly correlated with transcriptional activation [5,10]. Although the precise mechanisms by which histone acetylation alters transcription are poorly understood [9-12], there is tremendous pressure to understand these mechanisms, as impaired histone modification is linked to many disease states [13].The study of HAT and HDAC recruitment to promoters and their interaction with activators and repressors are essential for a better understanding of gene regulation. HATs and HDACs modi