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Figure 3 (A) Artistic representation of the ChIP-seq experimental outline. (B?C) Bar-plots showing the genomic distribution of high-confidence BCL9 peaks (B, 5303 total) and TBX3 peaks (C, 2369 total). (D) Overlap of the high-confidence peak groups between BCL9 and TBX3. (E) Selected result entries from motif analysis performed on the BCL9-TBX3 overlapping high-confidence peaks. Significant enrichment was found for TCF/LEF and Homeobox motifs. No TBX consensus sequence was detected in this analysis. (F) Select genomic tracks demonstrating occupancy of BCL9 and TBX3 within the Wnt Responsive Element (WRE) of known Wnt-target genes (Axin2, Ccnd1, Nkd1 and Lef1) and genes important in limb morphogenesis (Hand1 and Hand2). The scale of peak enrichment is indicated in the top-left corner of each group of tracks. In light blue the BCL9 (Salazar et al., 2019) and in orange the TBX3 replicates, and in green the control track (IgG). Genomic tracks are adapted for this figure upon visualization with IGV Integrative Genomic Viewer (https://igv.org/). Two independent replicates for BCL9 and TBX3 ChIP-seq experiments are shown. (G) Volcano plot displays all the differentially expressed genes (DEGs) in developing forelimbs, upon mutation of Bcl9/9l (Bcl9/9l-?1/?2 vs CTRL). DEGs were a total of 1143 (p<0.05), with 606 up-regulated and 537 down-regulated. N = 3 of individual mouse embryos for each condition were used for this analysis. (H) A significant portion (28.4%) of DEGs exhibited overlap with TBX3 ChIP-seq peaks. The overlap with TBX3 ChIP-seq peaks appeared statistically significant, in particular, when the down-regulated genes were considered. Hierarchical clustering of samples (3 CTRL versus 3 Bcl9/9l-?1/?2, right panel) based on genes overlapping between DEGs and genes annotated for TBX3 ChIP-seq peaks (normalized RNA-seq read counts, Ward?s clustering method, Euclidian distance). Annotation added for genes associated by Gene Ontology to Wnt signaling (Fgf10, Ptk7, Kremen1, Zfp703, Bmp2 and Gli3) and genes known as regulators of limb development (Meis2, Irx3 and Eya2).