Cell culture

All cells were cultured with penicillin (100 μ/ml) and streptomycin (100 μ/ml) and were regularly STR-verified and checked for mycoplasma by handing in samples prepared according to instructions at GATC Biotech (Konstanz, Germany). Primary human esophageal epithelial cells transformed with hTERT (EPC2-hTERT) (gift of K.K. Krishnadath)⁶⁴, were cultured with Keratinocyte SFM medium, supplemented with bovine pituitary extract at 50 µg/ml and EGF at 1 ng/ml (Thermo-Fisher Scientific). HET1A, the primary immortalized human squamous esophageal cell line Het-1A was a gift of J.W.P.M. van Baal (University Utrecht, The Netherlands). These cells were grown in EPM2 medium obtained from AthenaES, (Baltimore, Maryland, USA). The primary immortalized human BE cell line (BAR-T) was a gift of dr. J.W.P.M. van Baal who had, in turn, received them from dr. R.F. Souza (University of Texas Southwestern Medical Center, USA). These cells were grown in supplemented keratinocyte basal medium (KBM2), bought from Lonza (Basel, Switzerland), according to the method of Jaiswal et al.⁶⁵. KH2 mESCs were a gift of J. Gribnau and maintained in DMEM with 10% FCS, Non-Essential Amino Acids, sodium pyruvate, LIF, and β-mercapto-ethanol (embryonic stem cell medium; Supplementary Table ³). Dishes were coated with attachment factor protein solution (Thermo-Fisher Scientific). Irradiated mouse embryonic fibroblasts (3T3-Swiss albino cells (gift of J.W.P.M. van Baal), cultured in DMEM with 10% FCS, were used as feeder cells. HEK293T cells were cultured in DMEM with 10% FCS.