Fig 1: Binding of RUNX1 to evolutionary conserved RUNX1 heptamer motifs at the initiation site of TCRß rearrangements. (a) Runx1 expression in the lymphocytes of the outer cortex and the medulla (IHC; Scale bar: 100 µm). (b) Runx1 ChIP analyzed by real-time DNA PCR. The CD19 promoter was used as negative control and the Rag1 core promoter, which harbors two adjacent Runx1 binding sites, as a positive control. ChIP enrichment was calculated as fold change relative to input DNA with error bars representing the standard deviation of three biological replicates. Input and the y-axis were set to 1. Runx1 binding was detected for the Rag1 core promoter and the TCRß D1 and D2 segments. (c) Model of Runx1 binding to the Runx1 heptamer motifs of the murine TCRß D1 and D2 segments. (d) Alignment of TCRß D1 and D2 segments and their adjacent RSS heptamers show that RUNX1 binding sites (indicated by squares) are highly conserved at the 5' ends of D1 and D2 segments in Mammalia and Eutheria, respectively. At the 3' borders of TCRß D1 and D2 segments, RUNX1 heptamer motifs were found in approximately 45% of the analyzed mammalian species. In zebrafish (Danio rerio) used as outgroup only the TCRß D1 segment and no D2 segment is present. Previously reported C-FOS binding motifs at the 3' ends of the TCRß D1 and D2 segments are bold and underlined28.