Effect of HIPK2 on β-catenin-induced VEGF transcription. (A) Transcriptional activation of reporter plasmid expressing luciferase under the optimal TCF-responsive element (TOPFlash) was evaluated following co-transfection of H1299 cells with β-catenin and HIPK2 expression vectors. Thirty-six h after transfection, cells were assayed for luciferase activity. Results are expressed as Relative Luciferase Units (RLU) and represent the mean ± SD from three independent experiments performed in duplicate. *p < 0.001 versus β-catenin alone. (B, upper panel) H1299 cells were co-transfected with VEGF-luc reporter construct along with β-catenin and HIPK2 or K221R mutant, for detection of luciferase activity as above. The data shown as fold of luciferase activity represent the mean ± SD from three independent experiments performed in duplicate. *p < 0.001. (B, lower panel) VEGF mRNA levels were determined by RT-PCR analysis. GAPDH was used as loading control. One representative experiment from three independent experiments was shown. Densitometric analysis of VEGF levels was performed and normalized values to GAPDH mRNA levels were indicated.