Fkbp4

Total proteins were extracted using 1X cell lysing buffer (Cell Signaling Technology). Samples were separated by electrophoresis in SDS- polyacrylamide gels (12%) and transferred to PVDF membranes. Membranes were blocked for 1 h with PBST (PBS plus 0.1% Tween-20) containing 5% non-fat milk. Blots were incubated overnight at 4°C with primary antibodies in PBST containing 5% BSA at the manufacturer's recommended dilution. The antibodies were purchased from the following suppliers: FKBP4 (ProteinTech); anti-pS473 Akt, anti-pT308 Akt, anti-Akt, anti-pT286 Cyclin D1, anti-Cyclin D1, anti-E2F-1, anti-pT37/46 4E-BP1, anti-pS9 GSK-3β, anti-GSK-3β, and anti-mTOR (Cell Signaling Technology); anti-GAPDH and anti-Actin (Santa Cruz Biotechnology); anti-Tubulin and anti-PIK3R2 (Sigma-Aldrich). After washing, blots were incubated with either an anti-rabbit HRP, or an anti-mouse HRP-conjugated antibody (Cell Signaling Technology) for 1 h at 25°C. After washing, blots were developed with Supersignal WEST Pico Plus (Life Technologies SAS).

Antibodies were used in the following dilutions: NRF2 (1:1,000, Proteintech, #16396-1-AP), FKBP4 (1:1,000, Proteintech, #10655-1-AP), NR3C1 (1:1,000, Proteintech, #24050-1-AP), P62 (1:1,000, MBL, #PM045), Histone (1:1,000, CST, #3638), GAPDH (1:1,000, Proteintech, #60004-1-Ig), Flag (1:1,000, Sigma, #F3165), HA (1:1,000, Biolegend, #901514), Secondary antibody goat anti-mouse (1:2,500, HuaBio, #HA1006), secondary antibody goat anti-rabbit (1:2,500, HuaBio, #HA1001), Anti-RABBIT IgG (H&L) (GOAT) Antibody Rhodamine Conjugated (1:200, MULTISCIENCES, #RK-611-1002), Anti-RABBIT IgG (H&L) (GOAT) Antibody ATTO 488 Conjugated (1:200, MULTISCIENCES, #RK-611-152-122S), Anti-Mouse CD11c, PE (1:20, MULTISCIENCES, #AM011C04), Anti-Mouse CD86 (B7-2), APC (AM08605). Naringenin was purchased from APExBIO (#N1370), and resolved in DMSO at 10 mM. BC cells were treated by medium with Naringenin for different points in time.