Fig. 3: FASN enhances mutp53 palmitoylation to suppress mutp53 ubiquitination and stabilize mutp53.
From: Lipogenic enzyme FASN promotes mutant p53 accumulation and gain-of-function through palmitoylation
a The palmitoylation inhibitor 2-BP reduced protein levels of mutp53, but not wtp53, in different cells. Cells were treated with the 2-BP for 24 h before Western blot analysis. ABE analysis of palmitoylation of ectopic (b) or endogenous (c) mutp53 and wtp53 in cells. In b, p53-/- HCT116 cells expressing ectopic wtp53 or different mutp53 were employed for ABE assays. -: control vectors. HAM: hydroxylamine. Palm p53: palmitoylated p53. d FASN knockdown (left) or inhibition by TVB-3166 (TVB; right) drastically reduced mutp53 palmitoylation in p53-/- HCT116 cells determined by ABE assays. e, Identification of mutp53 palmitoylation sites by ABE assays in p53-/- HCT116 cells expressing R175H mutp53 or its different CS (cysteine to serine) mutants. Right panel: cysteine residues of p53. In b-e, to enable the standardized comparison of the p53 palmitoylation levels across various samples, the input p53 levels were normalized by adding different amounts of protein samples for Western blot analysis. In d, Actin was used as the control for FASN only. MG132 treatment largely abolished the inhibitory effect of 2-BP (f), FASN KO or knockdown (g), and the 4CS mutation (h) on mutp53 protein levels in cells. 2-BP treatment (i), FASN KO or knockdown (j), and the 4CS mutation (k) promoted mutp53 ubiquitination in cells analyzed by ubiquitination assays. Ub: Ubiquitin. FASN KO (l) and the 4CS mutation (m) reduced mutp53 protein half-life (T1/2) in cells. Cells were treated with CHX for different hours before Western blot assays. Data represent mean ± SD (n = 3 independent experiments). In a–m, Western blot data represent three repeats with similar results. Source data are provided as a Source Data file.
