D0819 500ml

ZR751 cells (RRID CVCL_0588) were obtained from the Lannigan laboratory⁶⁰ and grown in RPMI (Sigma Aldrich #R8758500ml) supplemented with 10% heat‐inactivated FBS (Corning™ #35016CV), 0.002% insulin (Sigma Aldrich #11376497001) and 50 IU penicillin, 50 mg/mL streptomycin (Corning™ #MT30001CI). HEK 293T cells (RRID CVCL_0063) were obtained from the Lannigan laboratory⁶⁰ and grown in DMEM with high glucose, L‐glutamine, phenol red, but not sodium pyruvate (Sigma Aldrich D0819‐500ML), 5% FBS, 1% Pen/Strep, and 1% Sodium pyruvate (Sigma Aldrich S8636‐100ML). The cell culture incubator parameters were as follows: 37˚C, 95% relative humidity, and 5% CO₂ concentration. The antibodies used for ChIP‐seq were anti‐Erα (Santa Cruz Biotechnology sc‐543X), anti‐H3K4me1 (Abcam ab8895), anti‐SP1 (Abcam ab13370), and sheep anti‐rabbit IgG Dynabeads M‐280 (Invitrogen™ 11203D).

The murine macrophage cell line RAW 264.7 (TIB-71, ATCC) (originating from BALB/c mice cells transformed with Abelson leukaemia virus [43 (link)]) was used as a model for the induction of a pro-immune or anti-inflammatory response. For facility of passaging, cells were cultured normally as a semi-adherent culture in 10 cm² suspension culture dishes at 37 °C, 5% CO₂ and 80% humidity in 12 mL DMEM (#D0819-500ML, Sigma-Aldrich, Saint-Quentin-Fallavier, France) medium supplemented with 5% FBS 100 U/mL penicillin and 0.1 mg/mL streptomycin (#P0781-100ML, Sigma). Cells were passaged by gentle flushing of the culture medium to remove the cells, with cells then pelleted (200 g, 3 min, RT), resuspended in medium and counted (Section 2.2.1). Cells were re-seeded at 5 × 10⁶ cells/mL or 10 × 10⁶ cells/mL for 2 or 3 days of growth, respectively.
For experiments, RAW 264.7 cells were seeded the day before treatment at a density of 3 × 10⁴ cells per well (100µL medium volume) in regular adherent 96-well culture plates (#655180, Greiner Bio-One, Les Ulis, France) or 3 × 10⁵ cells per well (300µL medium volume) in 12-well plates (#665180, Greiner Bio-One, Les Ulis, France).