Fig 5

Wnt-induced association of MLLT10/AF10-DOT1L with and regulation of Wnt target genes in HEK293T cells. (A–F) ChIP assays in HEK293T cells uninduced or induced with Wnt3A conditioned media at 2 and 12 h using antibodies specific for TCF4 (A), β-catenin (B), MLLT10/AF10 (C), DOT1L (D), H3K79 di-methyl (E), and H3K79 tri-methyl (F). The immunoprecipitated DNA was analyzed by qPCR using primers specific for the c-MYC and ZCCHC12 loci as indicated. Results are presented as percent immunoprecipitated over input and are representative of three independent experiments. (G) Comparison of the corresponding expression pattern after siRNA suppression of MLLT10/AF10, DOT1L, BRG1, and p300 in the Wnt induced condition. Heatmap showing 1988 Wnt regulated transcripts after 9 h Wnt-induction (relative to uninduced sample (no Wnt)) in HEK293T cells with greater than 1.5-fold variation, and the comparison of the corresponding expression pattern after siRNA suppression of MLLT10/AF10, DOT1L, BRG1, and p300 in Wnt-induced condition (relative to 9 h Wnt induction). Red, upregulated after Wnt; green, downregulated after Wnt induction; grey, missing data. Western blot analysis of MLLT10/AF10, DOT1L, BRG1, p300, and Tubulin upon siRNA depletion of each gene as indicated. (H) Venn diagram depicting the comparison of Wnt-induced genes and genes downregulated after MLLT10/AF10, DOT1L, BRG1, or p300 suppression in HEK293T cells after Wnt induction.