Cell culture

All cell lines were obtained from the Tissue Culture Core at Baylor College of Medicine (BCM) in 2017 from the ATCC, with validation by short tandem repeat testing completed at the Cytogenetics and Cell Authentication Core at MD Anderson. Cells were cultured at 37°C in 5% CO₂ and were tested for Mycoplasma every 6 months. Cell lines after passage 20 were discarded. T47D (ATCC cat# HTB-133, RRID: CVCL_0553) and MCF7 (ATCC cat# HTB-22, RRID: CVCL_0031) cells were grown in RPMI-1640 with l-glutamine (Corning, cat# 10–040-CV) supplemented with 10% FBS (Sigma-Aldrich, cat# F8067), glucose at 4.5 g/L (Sigma, cat# G5767), 10 mmol/L HEPES (GenDEPOT, cat# CA011), 1 mmol/L sodium pyruvate (GenDEPOT, cat# CA017), and 50 μg/mL gentamycin (GenDEPOT, cat# CR003–001). For hormone deprivation, cells were plated in culture media overnight, washed with Dulbecco's PBS, and maintained in phenol red-free, RPMI media (Thermo Fisher Scientific, cat# 11835) containing 10% charcoal-stripped serum (Sigma, cat# F6765) as supplemented as above (CSS media). CSS media were changed every 2–3 days for one-to-two weeks. Water-soluble E2 was purchased from Sigma (E4389).

Stable cell lines expressing YFP, truncated ESR1-e6, ESR1-WT, ESR1-Y537S, ESR1-D538G, ESR1-e6>YAP1, ESR1-e6>PCDH11X, ESR1-e6>DAB2, ESR1-e6>GYG1, ESR1-e6>SOX9, ESR1-e6> ARNT2-e18, ESR1-e6>PCMT1 and ESR1-e6>ARID1B, ESR1-e6>ARNT2-e2, ESR1-e6>LPP, ESR1-e6>NCOA1, ESR1-e6>TCF12, ESR1-e6>CLINT1, ESR1-e6>GRIP1 and ESR1-e6>TNRC6B were previously described (7).