A. p53 and p21 protein, as measured by Western blot, in A549 cells with shRNA targeting luciferase or RPS19 and treated with DMSO, 50 μM A-3, or 20 μM TFP.
B. qPCR measuring expression of TP53, P21, GADD45A, NOXA, or MDM2 in A549 cells with shRNA targeting luciferase or RPS19 and treated with DMSO, A-3, or TFP. Student t-test, *p < 0.05 compared to DMSO control.
(C-D) CD34+ cells were transduced with RPS19 or control shRNA and selected for successfully transduced GFP+ cells.
C. qPCR measuring p21 mRNA in CD34+ cells treated with increasing doses of TFP. Student t-test, *p < 0.05.
D. Western blot measuring RPS19 and p53 proteins in CD34+ cells treated for increasing lengths of time with 10 μM TFP.
(E-F) CD34+ cells were infected with RPS19 shRNA and selected for GFP+ successfully transduced cells by FACS. Western blots of p53 and GAPDH were quantified by densitometry.
E. Cells were pre-treated with TFP for 15-30 minutes before treatment with 10 μM CHX for increasing lengths of time before lysates were collected for TP53 and GAPDH protein quantification.
F. Cells were pre-treated with TFP for 2 hours before treatment with 20 μM MG-132 for increasing lengths of time before lysates were collected for TP53 and GAPDH protein quantification. Two-way ANOVA, *p < 0.05.