Slc40a1

RT-qPCR and western blotting analyses were performed as previously described [10 (link)]. The RT-qPCR reagents used in the experiments were purchased from Takara (Shiga, Japan). The primer sequences used for RT-qPCR are listed in Table S2. The primary and secondary antibodies used for western blotting against E-cadherin (Cat No. 14472), N-cadherin (Cat No. 13116), vimentin (Cat No. 46173), SLC7A11 (Cat No. 98051), COX2 (Cat No. 12282), GPX4 (Cat No. 59735), JUND (Cat No. 5000), and GAPDH (Cat No. 5174) were purchased from CST, transferrin (Cat No. ab82411), SLC40A1 (Cat No. ab239583) were purchased from Abcam, and the dilution ratio was determined according to the manufacturer’s instructions. All the full and uncropped western blots are uploaded as ‘Supplemental Material’.

β-elemene (>98%) (#E4418) was purchased from LKT. Stock solution at 100 mg/ml was made in ethanol and stored at -4°C. Cetuximab (#33657) was purchased from MCE. Antibodies against GPX4 (#GR251529-34), HO-1 (#GR3187585-3), Glutaminase (#GR3299063-1), SLC40A1 (#GR215168-39), Transferrin (#GR3207592-9), SLC7A11 (#GR3235736-6) were purchased from Abcam. N-Cadherin (#13116S), E-Cadherin (#14472S), MMP9 (#13667S), snail (#3879T), slug (#9585T), and GAPDH (#2118S) antibodies were obtained from Cell Signaling Technology. Vimentin (#SA10106DB) was purchased from ABGENT. Ki67 (#66434-1-Ig) was purchased from Proteintech. Deferoxamine (#CS-4479), Ferrostain-1 (#HY-100579), Necrostain-1 (#HY-15760) were purchased from MCE. Z-VAD-FMK (#V116) was obtained from Sigma Aldrich. Liproxstatin-1 (#S7699) and RSL3 (#S8155) were purchased from Selleck Chemical.

A phenolmethylsulfonyl–fluoride-containing lysis buffer containing radioimmunoassay precipitation was used to extract the total protein from glioma cells (Servicebio, Wuhan, China). In order to determine the protein concentrations in the samples, bicinchoninic acid (BCA) protein assays were performed (Servicebio, Wuhan, China). Separating proteins with sodium dodecyl sulfate polyacrylamide gels (Thermo Fisher Scientific, Waltham, MA, USA) led to their transfer to polyvinylidene fluoride membranes. Using skimmed milk powder to block the membranes, primary antibodies containing SLC7A11 (Cat No. 26864-1-AP), COX2 (Cat No. 66351-1-Ig), GPX4 (Cat No. 67763-1-Ig), JUN (Cat No. 66313-1-Ig), and β-actin (Cat No. 81115-1-RR) were purchased from proteintech; SLC40A1 (Cat No. ab239583) was purchased from abcam, in a dilution of 1:1000, was added for 2 h at room temperature and kept overnight at 4 °C. In total, 2 h of incubation was conducted at room temperature with a secondary antibody (in a dilution of 1:2000) after washing three times. A high-sensitivity ECL exposure solution was added and developed using an imager. β-actin was used as the loading control to calculate the relative protein expression.