Fig. 7

FOXO activates the expression of USP11, thereby upregulating PTEN. a Lysates from DU145 cells expressing SIRT1 shRNA treated with 25 μM resveratrol for 16 h were immunoprecipitated (IP) with anti-FOXO1, and the resulting immunoprecipitates were subjected to immunoblotting (IB). b, c Lysates from DU145 cells expressing two independent shRNAs against SIRT1 (b) or FOXO1 (c) treated with 25 μM resveratrol for 16 h were subjected to IB. d Lysates from DU145 cells pre-treated with 25 μM resveratrol for 16 h followed by treatment with cycloheximide (CHX, 100 μg ml⁻¹) for the indicated times were subjected to IB. n = 3, p value was determined by ANOVA. e Lysates from DU145 cells expressing USP11 shRNA treated with 25 μM resveratrol for 16 h were subjected to IB. f Lysates from DU145 cells expressing USP11 shRNA and treated with 25 μM resveratrol (Resv.) for 16 h and 10 μM MG132 for the last 4 h before harvesting were immunoprecipitated with anti-PTEN, and the resulting immunoprecipitates were subjected to IB. g Luciferase reporter analysis of the USP11 promoter in NIH-3T3 cells expressing constitutive, active FOXO1^{T24A,T256A,S319A} or FOXO3^{T32A,S253A,S315A}. n = 3. h Immunofluorescence analysis of FOXO1 in PTEN^-/- HCT116 cells treated with 5 μM psammaplysene A (PsA) for 24 h. Scale bars, 10μm. i Lysates from PTEN^+/+ HCT116 cells treated with 5 μM PsA for 24 h were subjected to IB. j Immunohistochemical analysis of FOXO3 or FOXO1 and USP11 in human prostate (left, n = 37) and TNBC (right, n = 68) tumor samples. Scale bars, 50μm. Correlation between FOXO localization and USP11 protein levels was determined by the PASS Pearson Chi-Square test (right). Error bars represent ± SEM. p Value was determined by Student’s t test (*p < 0.05, **p < 0.01, ***p < 0.001). TNBC, triple-negative breast cancer