Fig. 3. Dsg3 depletion causes further induction of p53 expression and activity in response to stress signals.
a Western blotting of siRNA pre-treated NTERT cells with and without UVB irradiation for the indicated proteins with the quantitation shown on the right (n = 3 biologically independent samples, *p < 0.05). b Western blotting for p53/p21WAF1/CIP1 in cells treated with or without actinomycin D (Act D, 5 nM) and mitomycin C (MMC, 5 µg/ml) for 24 h, respectively. The quantitation data are shown on the right. Enhanced expression of p53/p21WAF1/CIP1 was shown in cells with Dsg3 knockdown and treated with drugs. c Mechanical stretching induced increased expression of p53 and p21WAF1/CIP1/ Bax in Dsg3 KD cells. The siRNA pre-treated cells were seeded at confluent density in BioFlex plates and then subjected to cyclic strain (TX-5000, 20% amplitude, 1 Hz) for 4 h the following day. Lysates were extracted either immediately after strain or 2 h and 24 h later, respectively, after transferring to a stationary state, along with static control cells. d Western blotting analysis for PCNA and cyclin A in siRNA treated cells with and without UV. e Western blots for the indicated proteins upstream of p53 as well as phospho-p53-S20 in siRNA pre-treated cells with and without UVB (30 mJ/cm2). f Qnatitation of 53BP1 nuclear staining (n = 10, mean ± S.D., **p < 0.01, ***p < 0.001). g Western blotting in siRNA transfected cells treated in the presence and absence of ATM inhibitor KU55933 (20 µM) and p38 MAPK inhibitor SB203580 (20 µM), respectively. All cells were exposed to UVB (30 mJ/cm2) in this case. Cells were treated with drugs 1 h before UV and were grown overnight before lysate extraction. h The expression of p53 in the same samples as phosphorylated ATM and CHK2 (the last two corrected for total protein) following the indicated drug treatments in Dsg3-depleted cells. The data are expressed as the band intensities in the Dsg3-depleted cells relative to the corresponding scrambled siRNA controls, which were normalized to 1