Initiating RNA polymerase II and TIPs as hallmarks of enhancer activity and tissue-specificity

Abstract

In past years, many efforts were invested to define epigenetic features associated with enhancers of transcription. We propose that both transcription initiation and the H3K4me3 histone modification are among the best hallmarks of active enhancers in several primary tissues and extend the concept of large transcription initiation platforms (TIPs).

Figure 1

RNAP II and H3K4me3 are present at putative enhancers and increase the tissue-specific and -restrictive expression pattern of associated genes. (A) Examples of cell specific and non-specific RNAP II-bound enhancers in four different primary cell types. We used published data (see text) to isolate putative enhancers based on the presence of H3K4me1/H3K4me3/Ser5P at intergenic regions (±5 kb from any known annotation) in DP T cells, ESC, macrophages and neurons. The isolated regions are indicated as blue boxes underneath the H3K4me1 tracks. Using this strategy, we isolated tissue-specific enhancers (highlighted in blue) such as at Tcf7 in DP, Sox2 in ESC, Irg1 in macrophages, Lsamp/Gap43 in neurons as well as a conserved ones such as at Vps8 (from left to right, highlighted by gray boxes). (B) Tissue-specificity of putative enhancer selections. We assessed the tissue-specificity of genes associated to putative enhancers (neighboring upstream and downstream) by calculating the ratio of their expression level as compared with that of all remaining genes for all tissues available on BioGPS. Therefore, the higher ratios indicate higher tissue-specificity. The rank indicates the first occurrence of a tissue of interest (see legends A–D) when sorting the tissue-ratios from highest to lowest. With the exception of macrophages, either both the rank, but at least the ratio, increased when comparing a H3K4me1⁺/H3K4me3⁺/Ser5P to a H3K4me1⁺/H3K4me3⁻/HAT enhancer selection criteria. (C) Tissue-selective expression of genes associated with putative enhancer selections. To determine the tissue-selective expression of selected genes, we calculated the median expression level within our tissue(s) of interest for each cell type and plotted the differential to that of all remaining available tissues in the BioGPS database. Thus, the higher differentials represent the more tissue-selective expression pattern of the genes. Again, with the exception of macrophages, the H3K4me1⁺/H3K4me3⁺/Ser5P selection criteria (orange) improves the tissue-selective gene expression patterns as compared with H3K4me1⁺/H3K4me3⁻/HAT (blue).

Figure 2

TIPs at both promoters and enhancers increase the tissue-specific and -restrictive expression pattern of associated genes. (A) Examples of cell specific and non-specific promoter TIPs in four different primary cell types. We used the enrichment (at least two peaks with a minimal enrichment threshold across an area >400 bp) of RNAP II at promoters (−2/+1 kb) to isolate TIPs (highlighted in blue). Using this strategy, we isolated tissue-specific TIPs at Satb1 in DP, Sall4 in ESC, Emr1 in macrophages, Caly in neurons as well as conserved ones such as at Opa3 (from left to right, highlighted by gray boxes). The CpG islands track (red) was obtained from the UCSC browser and is shown as a reference. (B) Comparison of the tissue-specific expression of genes associated to promoters and putative enhancers associated to IGRs (H3K4me1⁺/H3K4me3⁺/Ser5P or TIPs). The tissue-specificity was calculated as described in Figure 1B. The presence of TIPs increased the tissue-specificity for both promoters and putative enhancers across all four tissues. The tissue-ratio rank either remained unchanged as first or increased in the case of DP and ESC. (C) Comparison of the tissue-selective expression of genes associated to promoters and putative enhancers (H3K4me1⁺/H3K4me3⁺/Ser5P or TIPs) as described in Figure 1C. The presence of TIPs either at promoters (purple) or putative enhancers (red) increased the tissue-restrictive expression pattern as compared with their isolation via H3K4me1⁺/H3K4me3⁺/Ser5P (green and orange respectively).