TETs enhance RUNX1-mediated DNA demethylation. (A) Confirmation of RUNX1, TET2, and TET3 overexpression in mock vector–overexpressing (293T-mock), RUNX1-overexpressing (293T-RUNX1), TET2-overexpressing (293T-TET2), TET3-overexpressing (293T-TET3), TET2 and RUNX1–co-overexpressing (293T-TET2-RUNX1), and TET3 and RUNX1–co-overexpressing (293T-TET3-RUNX1) 293T cells by western blotting. Immunoblotting for GAPDH is internal control. (B) Confirmation of RUNX1 (left), TET2 (middle), and TET3 (right) overexpression by qRT-PCR. y-axis represents average log2 fold change (FC) compared with mock vector–transduced cells. The error bars were SD. The experiments were performed in 3 biological replicates. (C) Overlap of demethylated CpGs between TET2-overexpressing and TET2 + RUNX1–overexpressing cells (TET2 vs TET2 + RUNX1) (blue), between TET3-overexpressing and TET3 + RUNX1–overexpressing cells (TET3 vs TET3 + RUNX1) (green), and between mock- and RUNX1-overexpressing cells (mock vs RUNX1) (red). The size of each circle represents total number of the demethylated CpGs. (D) Distribution of enrichment scores for RUNX1-binding motif within ±5000 bp from demethylated CpGs in TET2- and RUNX1-overexpressing 293T cells (red line), TET3- and RUNX1-overexpressing 293T cells (blue line), and RUNX1-overexpressing 293T cells (gray line). x- and y-axes show distance from CpG (bp) and enrichment score of RUNX1-binding motif, respectively.