Cell Culture Conditions and Treatments

Cell lines were grown at 37°C in a humidified incubator containing 5% CO₂. HeLa cells (ATCC CCL‐2), U2OS (ATCC HTB‐96) and HEK293T (ATCC CRL‐3216) cells were maintained in Dulbecco's modified Eagle's medium/nutrient mixture F‐12 GlutaMAX™ (DMEM/F‐12; Thermo Fisher Scientific) supplemented with 10% foetal bovine serum. Fibroblast cells derived from a patient with homozygous p.Arg435* FBXL4 have been previously published (Bonnen et al, 2013 (link)) (Alsina et al, 2020 (link)) and were cultured in DMEM/F‐12 GlutaMAX™ with 20% FBS and 5 mg/ml penicillin and streptomycin (Thermo Fisher Scientific). All cell lines were regularly screened for mycoplasma contamination. Where indicated, cells were treated with cycloheximide (CHX; 100 μg/ml; 66‐81‐9), deferiprone (DFP; 1 mM; 379409), DMOG (0.5 mM; D3695) and echinomycin (10 nM; SML0477), which were purchased from Sigma. MLN4924 (0.5 μM; 85923S) was obtained from Cell Signaling Technology. MG132 (10 μM; 474787) was purchased from Merck.