Fig. 8: TRIM21 interacts with SLC7A11 and modulates ferroptosis in prostate cancer cells.
A Co-immunoprecipitation (Co-IP) analysis of TRIM21 and SLC7A11 interaction in PC3 and DU145 cells. SLC7A11 was immunoprecipitated, and TRIM21 was detected by Western blotting. Reverse Co-IP was performed by immunoprecipitating TRIM21 and detecting SLC7A11. B Mass spectrometry analysis of SLC7A11 immunoprecipitates from HEK293 cells overexpressing CCDC719-13. The table lists the proteins identified with their corresponding molecular weights (MW) and log2 fold changes (Log2Ratio). C GST pull-down assay showing that CCDC719-13 promotes the interaction between TRIM21 and SLC7A11. EV (empty vector) serves as a control. D Western blot analysis of SLC7A11 protein levels in PC3 cells treated with cycloheximide (CHX, 500 nM) to block protein synthesis. Cells were transfected with either control or ShTRIM21. Protein levels were assessed at the indicated time points (0, 3, 6, 9, 18 h) post-treatment. GAPDH served as a loading control. E Co-IP assay demonstrating the ubiquitination of SLC7A11 in HEK293 cells co-transfected with Flag-CCDC719-13 and HA-Ub, with or without ShTRIM21. Cells were treated with MG132 (10 µM) to inhibit proteasomal degradation. F Western blot analysis of SLC7A11 protein levels in PC3 cells transfected as indicated. Relative cell viability of PC3 (G) and DU145 (H) cells transfected. Colony formation assay of PC3 (I) and DU145 (J) cells transfected as indicated. Quantification of cell death in PC3 (K) and DU145 (L) cells transfected with the indicated vectors. Relative ferroptosis levels in PC3 and DU145 cells transfected as indicated were determined by 4-HNE (M, N), MDA (O, P), and ROS (Q, R) levels.
