Fig. 4

Ferroptosis inducers promoted protein lipoylation through suppressing FDX1 protein mitochondria proteases dependent degradation A. DLAT and DLST protein lipoylation (lip-DLAT and lip-DLST), FDX1 and LIAS protein level in MHCC-97 H cells treated with 10µM sorafenib (Sora) for 24 h were blotted with indicated antibodies, GAPDH was used as the loading control B. lip-DLAT, lip-DLST, FDX1 protein level in MHCC-97 H cells with FDX1 knocking down and Sora (10µM) treatment for 24 h were blotted with indicated antibodies C. FDX1 mRNA expression in MHCC-97 H cells treated with Sora or Era 24 h was determined by real-time RT-PCR. D. The effect of sorafenib on FDX1 protein stability in MHCC-97 H (left) or QBC939 (right) cells with Cycloheximide (CHX, 100 µg/ml) and indicated concentration of Sora 12 h treatment was analyzed by immunoblotting E. FDX1 protein level in MHCC-97 H cells with DMSO, 10µM MG132 or 100nM Baf-A1 treatment for 12 h was analyzed by immunoblotting. ATF4 and p62 were used as positive controls of MG132 and Baf-A1 respectively F. The effect of AFG3L2 knockdown on FDX1 expression was analyzed by western blotting after 48 h of siRNA transfection in MHCC-97 H (left) or QBC939 (right) cells G. Cell viability of MHCC-97 H cells with Elesclomol-Cu after AFG3L2 knockdown in MHCC-97 H cells was measured with CCK-8 assay