JCS -- Candi et al. 119 (22): 4617 Figure IG1

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Fig. 1. TAp63 isoforms bind and transactivate the IKK ${alpha}$ promoter. (A) Schematic structure of the IKK ${alpha}$ promoter showing the different putative p53-like, Ets-1 and GATA-3 responsive elements. (B) Luciferase assay analysis showing the ability of TAp63 ${alpha}$ to induce IKK ${alpha}$ expression. The ratios of IKK ${alpha}$ promoter to transcription factor are: IKK ${alpha}$ :TAp63 ${alpha}$ / ${Delta}$ Np63 ${alpha}$ , 1:0.5; IKK ${alpha}$ :Ets-1, 1:2; IKK ${alpha}$ :TAp63 ${alpha}$ / ${Delta}$ Np63 ${alpha}$ :Ets-1, 1:2:2. The luciferase assay shown was performed in Saos-2 cells, and similar results were also obtained in HEK293 cells (data not shown). Results are shown as mean ± s.d. of three independent experiments. (C) Luciferase assay analysis showing the ability of TAp63 and ${Delta}$ Np63 isoforms to transactivate IKK ${alpha}$ in Saos-2 cells. Results are similar to those obtained in Saos-2 Tet-on inducible cells in B. The ratio of IKK ${alpha}$ promoter to TAp63/ ${Delta}$ Np63 isoforms was 1:2. Results are shown as mean ± s.d. of three independent experiments. (D) ChIP showing the binding of p63 protein to the IKK ${alpha}$ promoter. Lane 1, marker; lane 2, specific antibody (anti-HA); lane 3, non-specific antibody (nsp); lane 4, input+Dox; lane 5, input–Dox, (representative result of two independent experiments is shown). (E) Real-time PCR performed on Dox (Tet-on) inducible Saos-2 clones. TAp63 isoforms already induce a significant increase of IKK ${alpha}$ RNA upon 24 hours of Dox treatment, whereas ${Delta}$ Np63 ${alpha}$ and p53 do not significantly change IKK ${alpha}$ RNA levels. Results are mean ± s.d. of three independent experiments.