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Figure 4


Fig. 4. Differential ability of p63-mutant proteins to transactivate Dlx3. (A) Schematic representation of p63 transcripts and mutations. (B-C) Transcriptional regulation of the Dlx3 promoter by TAp63 (B) and {Delta}Np63 (C) wild-type and mutant proteins. H1299 cells were cotransfected with the Dlx3 reporter plasmid and expression vectors for TAp63- and {Delta}Np63-mutant isoforms. The basal activity of the reporter was set to 1. Each histogram bar represents the mean of three independent transfections. Standard deviations are indicated. (D) Proteins were corroborated by western blot analysis with anti-p63 4A4 antibody (Santa Cruz). (E) The level of endogenous Dlx3 mRNA upon transfection with p63 mutants in Saos-2 cells. Top panel, lanes: 1, mock; 2, TAp63{alpha}; 3, TAp63ß; 4, TAp63{gamma}; 5, TAp63F518V-AEC; 6, TAp63FS-EEC; 7, TAp63E639X-SHFM; and 8, TAp63{Delta}AA-LMS. Bottom panel, lanes: 1, mock; 2, {Delta}Np63{alpha}; 3, {Delta}Np63ß; 4, {Delta}Np63{gamma}; 5, {Delta}Np63F518V-AEC; 6, {Delta}Np63FS-EEC; 7, {Delta}Np63E639X-SHFM; and 8, {Delta}Np63{Delta}AA-LMS. GAPDH was used for normalization. (F) The level of endogenous Dlx3 mRNA upon transfection with TAp63{alpha} and TAp63{gamma} in HaCaT cells. Lanes: 1, mock; 2, TAp63{alpha} 0.5 µg; 3, TAp63{alpha} 2 µg; 4, TAp63{alpha} 4 µg; 5, TAp63{gamma} 0.5 µg; 6, TAp63{gamma} 2 µg; 7, TAp63{gamma} 4 µg. Cyclophillin was used for normalization.





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