Extended Data Fig. 3: Motif discovery of enhancers activated by Mesp1.

a, Motif discovery performed separately in Mesp1 ChIP-seq peaks classified as early, constant or late peaks. p-values are calculated through a binomial test. b, Same pipeline as panel a but separating peaks as de novo versus primed Mesp1 peaks. Stars represent the enrichment of the CAAATGG motif in pioneer peaks in comparison to non-pioneer peaks through a two-tailored Z-test. Data shown represent two biologically independent replicates. c, Quantification of the occurrence of all forms of bHLH motifs in Mesp1 ChIP-seq peaks, showing the prevalence of CAAATG motif, most often with an extra G. *** The top 2 bHLH motifs were notably over-represented (p < 0.00001) through a two-tailored Z-test, with z = 17.5 for CAAATG and z = 5.9 for CAGATG. d, Representative genomic locus where ATAC-seq and H₃K₂₇Ac ChIP-seq peaks are upregulated 24 h following Mesp1 expression but which are not directly bound by Mesp1 (red boxes), suggesting that their regulation is mediated by other TFs, whose expression is induced directly or indirectly by Mesp1. Gata4 ChIP-seq data³¹ (green) shows a strong overlap between these de novo opened peaks not bound but induced by Mesp1. e, Motif discovery searching for known TF binding sites within ATAC-seq peaks that get opened by Mesp1 but are not directly bound by Mesp1. p-values are calculated through a binomial test. f, Quantification of peaks containing a Gata4 motif, both in Mesp1 ChIP-seq peaks and ATAC-seq peaks UP without Mesp1 binding, separated as peaks bound by Gata4 or not bound by Gata4 using a published ChIP-seq dataset³¹. *** Gata4 motifs are statistically notably enriched in Gata4-binding peaks in Mesp1 ChIP-seq (z = 8.34, p < 0.00001) and in ATAC-seq peaks UP (z = 13.6, p < 0.00001). Data shown represent two biologically independent replicates; n = 1 for previously published Gata4 ChIP-seq³¹. These values were calculated through a two-tailored Z-test.