Figure.4. SLC7A11 promoter mutation cells are susceptible to ferroptosis.

A. Schematic diagram of SLC7A11 promoter WT and mutant H1299 cells.

B. The protein levels of SLC7A11 in SLC7A11 promoter WT and mutant H1299 cells were analyzed by WB.

C. Relative mRNA expression levels of SLC7A11, GSS, GCLC and GPX4 in SLC7A11 promoter WT and mutant H1299 cells were analyzed by qRT-PCR, *P<0.05 (Student's t-test). SLC7A11 but not other ferroptosis related genes was decreased in SLC7A11 promoter mutation cells.

D. ChIP results of SOX2 binding on SLC7A11 promoter in WT and mutant H1299 cells. The mutation largely abolished SOX2’s binding on the SLC7A11 promoter.

E. Relative cysteine levels in SLC7A11 promoter WT and mutant H1299 cells were showed, *P<0.05 (Student's t-test).

F. Relative GSH levels in SLC7A11 promoter WT and mutant H1299 cells were showed, **P<0.01 (Student's t-test).

G. Lipid ROS levels of SLC7A11 promoter WT and mutation H1299 cells treated with 5 μM Erastin for 10 hours.

H. SLC7A11 promoter WT and mutant H1299 cells were treated with different dose of Erastin for 20 hours and cell viability of indicated cells was measured. The data was representative of three independent experiments.

I. SOX2 WT and KO H1299 cells were treated with different dose of Erastin for 20 hours and cell viability of indicated cells was measured. The data was representative of three independent experiments.

J. Indicated H1299 cells were treated with 10 μM Erastin for 20 hours, 1 μM Fer-1 and 50 μM DFO treatment could rescue Erastin-induced cell death. Cell viability of indicated cells was measured, ***P<0.001 (two-way ANOVA test).