Figure 3
CHIR99021 inhibits p53-dependent PUMA induction and p53 K120-acetylation.
Crypt culture were subjected to 5 Gy, or mock (Un) irradiation and harvested at indicated time points. (A) The expression of indicated proteins at 24 h in cultured crypts (3 wells pooled) was determined by western blotting. β-actin (Actin) was used as the control for loading. (B) Expression of PUMA and p21 transcripts was analyzed by real-time RT-PCR. (C) Cultured crypts were infected with an adenovirus encoding PUMA (Ad-PUMA) for 24 h and PUMA mRNA levels and protein (HA) were analyzed by real-time RT-PCR and western blotting. (D) Cultured crypts were infected by Ad-PUMA at the same time with or without CHIR99021 treatment. Left, quantitation of enteroids with 5 or more buds at day 4. Right, representative pictures of enteroids at day 4. Scale bar, 100 μm. Values are means ± SD, n = 3 wells from three different mice in each group. ***P < 0.001, **P < 0.01, *P < 0.05.
