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. 2004 Feb 1;18(3):261–277. doi: 10.1101/gad.1165804

Figure 2.

Figure 2.

ER stress prevents p53 stabilization and impairs p53-mediated apoptosis in response to genotoxic stress. (A) WI-38 cells were treated with 1 μM of ADR alone or with either 10 μg/mL of TM or 1 μM of TG for the indicated times. Protein extracts (50 μg) were used for immunoblotting with an anti-p53 rabbit polyclonal antibody. A nonspecific (NS) band on the same blot was used as loading control. (B) WI-38 cells were treated with 5 nM ACD in the absence or presence of either 1 μg/mL of TM or 0.1 μM of TG for the indicated times. Protein extracts (50 μg) were immunoblotted with an anti-p53 rabbit polyclonal antibody. Blot was stripped and reprobed with an anti-actin antibody. (C) HCT116 p53+/+ cells were treated with 375 μM of 5-FU in the absence or presence of either 1 μg/mL of TM or 0.1 μM of TG for the indicated times. The protein levels of p53 were detected by immunoblotting of 50 μg protein extracts with anti-p53 rabbit polyclonal antibody. A nonspecific band on the same blot was used as loading control. (D,E) HCT116 p53 +/+and HCT116 p53-/- cells were pretreated with either 0.3 μg/mL of TM or 0.03 μM of TG for 1 h, followed by treatment with 375 μM of 5-FU for 3 h. The cells were fed with fresh medium and subjected either to FACS analysis 48 h post-treatment (D) or colony formation assay (E) as described in Materials and Methods. The numbers represent the average values of colony formation quantification for each treatment from three independent experiments. Untreated (CON) cells were scored as 100%.