Fig. 3: HOX13 transcription factor proteins bind to the II1 enhancer in T-DOM.

a ATAC-Seq and CUT&RUN reads mapped to the II1 enhancer sequence, either in its native environment within C-DOM (left column; mm10 chr2:74,074,674-74,076,672), or its targeted recombination site within T-DOM (breeding line 542, right column; mm10 chr2:75,268,925-75,270,923). The II1 C-DOM element is not accessible by ATAC-Seq in E12.5 forebrain (FB), nor in proximal forelimb cells (PFL) samples. At E12.5 it becomes highly accessible in distal forelimb cells (DFL) and is strongly bound by HOX13 proteins. The II1 element in T-DOM has low accessibility in the FB and PFL samples, even though there is a high transcription of the transgene in PFL. Like the II1 element in C-DOM, the II1 enhancer in T-DOM is occupied by HOX13 proteins in distal limb cells. It also shows an additional peak over the HBB promoter (orange arrow). This peak is likely a non-specific signal resulting from promiscuous MNase activity used in the CUT&RUN technique. In all samples but HOXD13 in the II1 T-DOM allele, experiments were performed in duplicate. One biological replicate is plotted as a solid color and the other is shown as a superimposed black line (n = 2). Green rectangles below indicate the position of the II1 enhancer element relative to the peak signal; the position of the four HOX13 binding sites is indicated by pink lines. b On top is a schematic of the II1 enhancer element with the four HOX13 motifs indicated as pink bars. The pink bar with an asterisk indicates the motif position that is not near the HOX13 and ATAC peaks. At the bottom are the three HOX13 motifs identified by HOMER motif discovery in the CUT&RUN experiments here and the E11.5 whole forelimb ChIP-seq²⁵.