A method to build core regulatory circuitry. (A) Graphical description of the method used to create core regulatory circuitry (CRC) models. 1. Identification of SE-assigned expressed TFs. 2. Identification of the TFs that are predicted to bind their own SE, considered as auto-regulated. 3. CRCs are assembled as fully interconnected loops of auto-regulated TFs. (B) Cartoon showing: 1. TF-assigned SE constituents defined by H3K27ac ChIP-seq peak signals; 2. TFs having at least three DNA-binding sequence motif instances in their SE constituents are considered auto-regulated; 3. TFs with SEs having at least three DNA-binding sequence motif instances for each of the other predicted auto-regulated TFs together form an interconnected auto-regulatory loop. (C) Metagenes for the ChIP-seq signal for H3K27ac (left) and for the average ChIP-seq signal for POU5F1, SOX2, and NANOG (right) in H1 hESCs in the region ±5 kb around the center of the SE constituents. (D) Average percentage of DNA-binding motifs that are actually bound by the TFs from ChIP-seq data for POU5F1, SOX2, and NANOG in H1 hESCs, in either SE constituents or sets of random genomic sequences of the same size. (E) Venn diagram showing the average numbers, across 84 samples, of: 1. TFs having motifs that are expressed (445 TFs); 2. TFs having motifs that are expressed and assigned to a SE (61 TFs); 3. TFs having motifs that are expressed and assigned to a SE and that are predicted to bind their own SE (39 TFs); 4. TFs that are part of the CRC model (15 TFs).
