ACSL4/ELOC mediates the ferroptosis-inhibiting process of ROLLCSC in tumor cells. (IMAGE)
Caption
(A) Venn diagram showing methylation-related proteins specifically interacting with the sense strand. (B) RIP verified the direct binding of anti-ACSL4 and ROLLCSC, and anti-IgG served as a negative control. (C) WB experiments identified the direct binding of anti-ACSL4 and ROLLCSC. (D) WB experiments identified the direct binding of anti-ACSL4 and ROLLCSC fragments. (E) Western Blot was employed to examine the influence of ROLLCSC on the protein level of ACSL4 subsequent to MG132 (1 μM) and CHX (1 μM) treatment. (F) Western blot of the total ubiquitination proteins and ACSL4 from control and ROLLCSC overexpression of lentivirus transfection LLC cells. (G) Immunoprecipitation and mass spectrometry were used to screen the anti-ACSL4 interacting proteins in LLC-oeROLLCSC cells. (H) Western blot confirmed the direct binding of ACSL4 to ELOC in LLC-oeROLLCSC cells. (I) Western blot was used to detect the protein expression of ACSL4 after transfection of si-Eloc in LLC-oeROLLCSC cells. (J) Western blot was used to detect the expression of ACSL4 protein and total ubiquitin protein after transfection of si-Eloc in LLC-oeROLLCSC cells. (K) Investigated the effect of Acsl4 overexpression on MDA and ROS levels in LLC-oeROLLCSC cells. (L) Investigated the effect of Eloc knockdown on MDA and ROS levels in LLC-oeROLLCSC cells. (M) Liperfluo reagent was used to verify the effect of Acsl4 overexpression and Eloc knockdown on intracellular lipid peroxidation levels in LLC-oeROLLCSC cells; Red = intracellular lipid peroxidation, DAPI = nucleus. (N) Representative images of transwell results suggest the effect of Acsl4 and Eloc on the migration (upper) and invasion (down) ability of LLC cells. (O, P) Lung metastatic lesions were assessed by fluorescence imaging and H&E staining after euthanasia. Images are representative of three independent mice (n = 3). (Q) Investigated the effect of Acsl4 overexpression on ROS and MDA levels in lung metastases of C57BL/6 mice. (R) Investigated the effect of Eloc knockdown on ROS and MDA levels in lung metastases of C57BL/6 mice. (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. ns, not significant. The results represent three independent experiments.)
Credit
Yu-Han Zhang, Jia-Cheng Xie, Ting Ye, Shi-Meng Guo, Xue Han, Si Yang, Lei Shi, Yi-Shi Li, H. Rosie Xing, Jing-Yu Li, Jian-Yu Wang
Usage Restrictions
Credit must be given to the creator. Only noncommercial uses of the work are permitted. No derivatives or adaptations of the work are permitted.
License
CC BY-NC-ND