Fig. 2: Doxo and Epi induce disaggregation of p63 L514F protein in H1299 cells. | Cell Death Discovery

Fig. 2: Doxo and Epi induce disaggregation of p63 L514F protein in H1299 cells.

From: Anthracyclines disaggregate and restore mutant p63 function: a potential therapeutic approach for AEC syndrome

Fig. 2

A SDS-PAGE followed by Western blot for p63 in H1299 cells transfected with wild-type p63 and mutant L514F. 24 h after transfection, cells were treated with 100 μg/mL CHX and incubated with 2 μM Doxo or 1 μM Epi. At the indicated time points, cells were harvested, lysed, and analyzed for p63 levels. The asterisk indicates a non-specific band. Vinculin levels were measured as a loading control. B Quantitative analysis of the p63 protein levels in cells after CHX treatment. Data are represented as mean ± SD (n = 3). C SDS-PAGE followed by Western blot for p63 in H1299 cells transfected with wild-type p63 or mutant L514F. 24 h after transfection, cells were treated with 100 μg/mL CHX, a combination of 100 μg/mL CHX and 10 μM MG132 to inhibit the proteasome, or a combination of 100 μg/mL CHX and 100 nM Concanamycin A to inhibit autophagy. Cells were incubated with 2 μM Doxo or 1 μM Epi for 24 h. Cells were harvested and analyzed for p63 levels. As controls of treatment efficiency, LC3 A/B-I and II protein level increased when autophagy was inhibited, and antibodies anti-Ubiquitin revealed an accumulation of poly-ubiquitinated proteins when the proteasome was blocked. Vinculin levels were measured as a loading control. D Blue-native (BN) PAGE followed by Western blot for p63 in H1299 cells transfected with mutant L514F alone or together with wild-type p63 and treated with 2 μM Doxo or 1 μM Epi for 48 hr. Samples were normalized for p63 amount by Western blot analysis. m monomer, d dimer, t tetramer.

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