Psl01

SW620, SW837, SW480, DLD1, and HEK293T cells were purchased from the ATCC. Wild-type and CBS KO MEFs were generated from day 13.5 embryos isolated from wild-type and CBS KO mice (C57B6, #002461, The Jackson Laboratory). DLD1 cells were cultured in RPMI-1640 medium (Shanghai Basal Media Technologies Company, L210KJ). HEK293T cells and MEFs were cultured in DMEM (Shanghai Basal Media Technologies Company, L110KJ). SW480, SW620, and SW837 cells were cultured in L-15 medium (Shanghai Basal Media Technologies Company, L620KJ). For cystine restriction experiments, cells were grown in L-cystine-free RPMI 1640 medium or in medium supplemented with 65 mg/L L-cystine. All cells were grown in a medium containing 10% fetal bovine serum (FBS; A24G00J, Gemini) supplemented with 100 units/mL penicillin and streptomycin (PSL01, Caisson). All cells were incubated at 37 °C in a humidified atmosphere containing 5% CO₂. All cell lines were identified using short tandem repeat (STR) profiling.

Cell lines OVCAR10, OVCAR5, SKOV3 and RMG1 were cultured in Medium 199 (M5017, Sigma-Aldrich,) and MCDB 105 (M6395, Sigma-Aldrich,) at 1:1 ratio with 5% fetal bovine serum (F0926, Sigma-Aldrich,) and 1% streptomycin/pencillin (PSL01, Caisson Labs, Smithfield, UT, USA). Cell lines OVSAHO, OVCAR8 and IGROV1 were cultured in RPMI (RPL03, Caisson Labs) with 5% fetal bovine serum and 1% streptomycin/penicillin. PERK-MEF^−/− (CRL-2976, ATCC, Manassas, VA, USA) and GCN2-MEF^−/− (CRL-2978, ATCC) were cultured in DMEM, 0.1 mM non-essential amino acids (MT25025CL, Fisher, Waltham, MA, USA), 0.05 nM 2-mercaptoethanol (M3148, Sigma-Aldrich), 10% fetal bovine serum, and 1% streptomycin/penicillin. MDA-MB-241 cells were cultured in RPMI, 0.1 mM non-essential amino acid with 5% fetal bovine serum and 1% streptomycin/penicillin. All cell lines used in this study were cultured in a 37 °C humidified incubator with 5% CO₂ and were periodically checked for mycoplasma contamination. MEF cells were purchased directly from the vendor for this study. The identity of all remaining cell lines was confirmed with STR genotyping.