Cell Culture Protocols for HEK293T and U87 Cells

HEK293T (Thermo Fisher Scientific) and U87 cells (American Type Culture Collection) were cultured in Dulbecco's modified Eagle medium (DMEM; Gibco, 41965039) supplemented with 10% foetal bovine serum (FBS; Gibco, 10270106), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco, 5070063). 4EHP-KO and GIGYF2-KO HEK293 cells have been described previously (Xu et al., 2022 (link); Zhang et al., 2021 (link)). Parental, 4EHP-KO and GIGYF2-KO HEK293 cells were maintained in DMEM supplemented with 10% FBS, 1% penicillin/streptomycin, 100 µg/ml zeocin (Invitrogen, 460509) and 15 µg/ml blasticidin (BioShop, BLA477). All cells were cultured at 37°C in a humidified atmosphere with 5% CO₂ and regularly tested for the presence of mycoplasma contamination using a mycoplasma detection kit (Applied Biological Materials, G238).