To test whether CAGE expression can identify cell type-specific enhancer usage in vivo, ChIP-seq (H3K27ac and H3K4me1), DNA methylation and triplicate CAGE analyses were performed in five primary blood cell types, and compared to published DHS data (www.roadmapepigenomics.org, Supplementary Table 4). CAGE-defined enhancers were strongly supported by proximal H3K4me1/H3K27ac peaks (71%) and DHSs (87%) from the same cell type. Conversely, H3K4me1 and H3K27ac supported only 24% of DHSs distal to promoters and exons and only 4% of DHSs overlapped CAGE-defined enhancers (Supplementary Fig. 15), suggesting that a minority of promoter-distal DHSs identify enhancers. From the opposite perspective, only 11% of H3K4me1/H3K27ac loci overlapped CAGE-defined enhancers and untranscribed loci showed weaker ChIP-seq signals than transcribed ones (Supplementary Fig. 16). Moreover, there was a clear correlation between CAGE, DNase I hypersensitivity, H3K4me1 and H3K27ac for CAGE-defined enhancers expressed in blood cells (Fig. 3a). Accordingly, cell type-specific enhancer expression corresponds to cell type-specific histone modifications (Fig. 3b). The majority of selected cell type-specific enhancers could be validated in corresponding cell lines and were associated with cell type-specific DNA demethylation (Supplementary Text, Supplementary Fig.
