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. Author manuscript; available in PMC: 2013 Aug 7.
Published in final edited form as: Oncogene. 2012 Mar 5;32(3):388–396. doi: 10.1038/onc.2012.63

Figure 5. Regulation of p53 activity by knocking down endogenous RPS14 or RPS19.

Figure 5

(A) and (B) RPS14 knockdown activates p53 but also inhibits ribosomal stress-induced p53 activation. HCT116p53+/+ cells (A) and A549 cells (B) were transfected with scrambled siRNAs or RPS14 siRNAs. Cells were treated with 5 nM actinomycin D (Act D) or 10 µM mycophenolic acid (MPA) before harvesting for IB with antibodies as indicated. (C) RPS14 knockdown results in RPL5- and RPL11-dependent p53 activation. A549 cells were transfected with combinations of scrambled siRNAs, RPS14 siRNAs, RPL5 siRNAs or RPL11 siRNAs. Expression of p53, p21, RPS14, RPL11 and β-actin were detected by IB using antibodies as indicated. Messenger RNA levels of RPL5 and GAPDH were detected by RT-PCR. (D) Knockdown of RPS19 results in RPL5- and RPL11-dependent p53 activation. The same experiment as shown in panel C was conducted except the RPS19 siRNAs were used here. Expression of p53, MDM2, RPS19, RPL11 and β-actin were detected by IB using antibodies as indicated. Messenger RNA levels of RPL5 and GAPDH were detected by RT-PCR. (E) (F) Knockdown of RPS14 causes p53-dependent cell cycle arrest. HCT116p53+/+ cells (E) and HCT116p53−/− cells (F) were transfected with scrambled siRNAs or RPS14 siRNAs for 48 h, followed by FACS analysis. The mean percentages of cells in each phase obtained from three separate experiments are presented. Bars indicate standard deviations. “*” indicates P<0.05. (G) A schematic model for p53 activation by general ribosomal stress (upper panel) or 40S subunit perturbation (lower panel).