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. 2015 Dec 4;15(1):52–63. doi: 10.1080/15384101.2015.1120927

Figure 1.

Figure 1.

Gamma-irradiation induces p53 Ser15 phosphorylation and nuclear accumulation. (A) Immunofluorescent staining of mESCs with antibody to phospho-p53 (Ser15) 2 h post-irradiation (6 Gy) (green). Nuclei were stained with To-Pro3 (blue). Scale bar 10 μM. (B) Western blotting analysis of protein extracts from non-irradiated and irradiated mESCs. Blots were stained 2, 4 and 8 h post-irradiation (6 Gy) using antibody to phospho-p53 (Ser15), acetyl-p53 (Lys 379) and total p53. (C) A luciferase assay of control and irradiated mESCs after transient transfection with luciferase reporter plasmid PG13-Luc. At time points 4 and 24 h post-irradiation cells were subjected to luciferase activity assays. Pifithrin-α, an inhibitor p53-transcriptional activity (10 μM), was added to the 4 h reaction as a negative control. Error bars correspond to SEM calculated for 3 replicates. (D) Western blotting analysis of control, transfected with luciferase reporter plasmid PG13-Luc and irradiated with 3 Gy (4 h post-irradiation) mESCs. Irradiated cells were taken as control. Blots were stained using antibody to phospho-histone H2AX (Ser139) (γH2AX), phospho-p53 (Ser15) and total p53.