Cd2+ triggers nuclear translocation of TCF4 and activates TCF4/β-catenin mediated transcription of cell proliferation and survival genes in kidney PTC. (A) Distribution of TCF4 protein in cytoplasmic and nuclear fractions of WKPT-0293 Cl.2 cells ± Cd2+ was determined by immunoblotting. (B) Increased association of immunoprecipitated β-catenin with TCF4 by Cd2+ in WKPT-0293 Cl.2 cells. (C) Cd2+ induces TCF transcriptional activity. Luciferase activity of TOPflash or FOPflash transfected cells treated with 25 μM Cd2+ for 3-12 h. Data were corrected for protein and normalized to controls at each time point. Means ± SEM (n = 3-8) are shown. Student's unpaired t-test compares Cd2+ treated cells to respective controls. (D) Effect of Cd2+ exposure on mRNA of Wnt pathway target genes. GAPDH was used as housekeeping gene. (E) EMSA of TCF4 binding to the ABCB1 promoter region. Extracts from controls (ctl), Cd2+ (25 μM) treated cells, cells exposed to 3 h Cd2+ followed by incubation with a 200-fold excess of competing unlabeled oligonucleotides (comp.), and lysate-free sample (blank) were loaded. Binding of oligonucleotides to TCF4 was enhanced upon Cd2+ exposure. (F) Effect of TCF4 overexpression on Cd2+ induced c-Myc mRNA expression. WKPT-0293 Cl.2 cells transiently transfected with full length human TCF4 (TCF4), human TCF4 lacking the interaction domain for β-catenin (ΔN-TCF4), or empty vector were treated with ± 25 μM Cd2+. TCF4 overexpression enhanced basal and Cd2+-induced c-Myc expression, whereas ΔN-TCF4 had no effect.