To refine our search and identify regions in the promoter where TF binding may affect H4K5ac occupancy, we profiled the coverage of H4K5ac on all genes, on genes with a TFBS at 500 bp, 800 bp or 1100 bp upstream Nilotinib of the TSS, and on genes with no TFBS 100 bp upstream of the TSS. Using the average coverage of H4K5ac of all genes as baseline, we observed that the presence of a TFBS at position ?500 bp or ?800 bp, and ?1100 bp resulted in modest a reduction in H4K5ac relative to baseline coverage at that position. However, genes with no TFBS upstream of 100 bp resulted in significantly higher H4K5ac in both the promoter and CDS, approxi mately 1 kb relative to the TSS. Based on the increase of H4K5ac coverage in the ab sence of a TFBS upstream of 100 bp, we focused our analysis in this region, proximal to the TSS.

We com Inhibitors,Modulators,Libraries pared the contribution of acetylated gene clusters in the presence or absence of a TFBS relative to 150 bp of the TSS either no TFBS present in the promoter or no TFBS, one TFBS, or mul tiple TFBS 150 bp upstream of the TSS. Gene clusters with relatively no enrichment for H4K5ac or H4K12ac constituted a larger proportion of genes regardless of whether a TFBS was present or not. However, in the presence of at least one TFBS within 150 bp of the TSS, the contribution of cluster 4 for H4K5ac in FC, cluster 3 for H4K5ac in control, and cluster 1 for H4K12ac after CFC increased from approximately 10% to 20%, compared to the same clusters when no TFBS was present. To a lesser extent, cluster contribution was also increased in the presence of one TFBS 150 bp upstream of the TSS, but was diminished in the presence of multiple TFBS.

These observations provide novel insight into H4K5ac mediated regulation of gene transcription and support the notion that Inhibitors,Modulators,Libraries TF binding and acetylation are mutually exclusive in the promoter. However, H4K5ac is increased when Inhibitors,Modulators,Libraries TF binding occurs prox imal to the TSS. The observed increase in acetylation and transcription at proximal TFBS may be attributed to the recruitment of transcriptional machinery including TFs and RNA polymerase II, which is also known to occupy positions near the TSS in actively transcribed genes. Add itionally, recent ENCODE studies have shown that a set of TFs is strongly associated to positions proximal to the TSS and that transcriptional initiation is determined by stereotyped TF binding in this region, approximately Inhibitors,Modulators,Libraries 100 to 200 bp upstream of the TSS.

Acetylated nucleosomes further away in the promoter, Inhibitors,Modulators,Libraries greater than 1 kb from the TSS, may either be more strongly bound and less easily displaced by TF binding, or former they may be regulatory regions which do not depend on the presence or acetylation of nucleosomes. As expected, IgG IP control clusters were uniformly proportioned in the presence or absence of a TFBS.