Mda mb 231

Nonmalignant (MCF10A, MMuMG) and malignant (MDA‐MB‐468, MCF‐7 and MDA‐MB‐231) breast cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). Nonmalignant breast cell lines were cultured in mammary epithelial cell growth medium with supplements (Lonza, Walkersville, MD, USA), and malignant breast cell lines were cultured in RPMI 1640 medium (Thermo Scientific, Rockford, IL, USA) supplemented with 10% FBS (Gibco, Gaithersburg, MD, USA) at 37 °C in an atmosphere of 95% air and 5% CO₂.
Stable cell lines of MDA‐MB‐231 were generated by integration of retroviral shRNA vectors specific for GHR or a control vector from OriGene (Rockville, MD, USA).

Jurkat leukemia T cells and human breast cancer lines MDA-MB-231 and MDA-MB-468 were purchased from ATCC (Manassas, VA, USA). The cell lines were authenticated by the ATCC using short tandem repeat profiling and passaged in our laboratory for fewer than 6 months after receipt. Jurkat leukemia T cells were cultured in RPMI-1640 (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% FCS at 37°C in 5% CO₂. MDA-MB-231 and MDA-MB-468 cells were cultured in Dulbecco's modified Eagle's medium (Sigma-Aldrich) supplemented with 10% FCS at 37°C in 5% CO₂. Short hairpin RNA for human ERO1-α (TR313168) was purchased from OriGene (Rockville, MD, USA) and transfected to MDA-MB-231 cells using Lipofectamine 2000 (Life Technologies). To establish ERO1-α-overexpressing cells, MDA-MB-231 cells were transfected with pIRES puro3 myc2/ERO1 or an empty vector as a control using Lipofectamine 2000 (Life Technologies) per the manufacturer's instructions. Cells were stably propagated under puromycin selection (2 μg/ml). siRNA to ERO1-α and control siRNA were purchased from Origene and transfected to MDA-MB-468 cells. Four days after transfection, cells were harvested and used in flowcytometric assay and gene expression assay. SiRNA to HIF-1α and control siRNA were purchased from Origene and transfected to MDA-MB-231 cells.

Two breast cancer cell lines T47D and MDA-MB-231 were purchased from ATCC, and cultured in DMEM medium. To stably knock down RUNX1-IT1, two shRNAs targeting RUNX1-IT1 (#1: 5′-TCGAAGACATCGGCAGAAA-3′; #2: ACCACTCCACTGCCTTTAA; Negative control (NC): TTCTCCGAACGTGTCACGT) were inserted into pLKO.1-puro lentiviral vector, followed by infection into T47D and MDA-MB-231 cells. The stable clone was screened by puromycin for 1 week. GPX4 vector was purchased from OriGene (#RC208065) and transfected into T47D and MDA-MB-231 cells to overexpress GPX4. Besides, the full-length coding sequences (CDS) of IGF2BP1 and deletion of KH3/4 mutant were cloned into pEGFP-N1 vector, followed by transfection into HEK293T cells to overexpress IGF2BP1 and its mutant. Protein affinity purification was conducted by using Anti-GFP Magnetic Beads (#AE079, ABclonal, MA, USA) as per the manufacturer’s instructions. Cell transfection was carried out using Lipofectamine 3000 (Thermo Fisher Scientific, CA, USA).