Fig. 5

TC2N promotes the degradation of FASN via blocking the interaction between PTEN and FASN. A The extracts from M231 cells with control lentivirus or TC2N overexpression lentivirus treated with 20 µM cycloheximide (CHX) for the indicated times were subjected to WB (Upper). Relative FASN protein levels were quantified by ImageJ software (Lower). B M231 cells with control lentivirus or TC2N overexpression lentivirus were transfected with 2 µg of ubiquitin-expressing plasmids. At 24 h after the transfection, cells were treated with 20 µM MG132 for 24 h. Cell extracts were subjected to IP with anti-FASN antibody and further analyzed by WB with anti-Ubiquitin and anti- Tyrosine antibody. Normal IgG was used as a negative control. Whole-cell lysates were used as a positive control (Input). C The extracts from BC cells were subjected to IP with anti-Flag antibody and further analyzed by WB with TC2N, FASN, TRIM21 and PTEN antibodies. Normal IgG was used as a negative control. Whole-cell lysates were used as a positive control (Input). D and E BC cells were treated with 20 µM MG132 for 24 h, and then cell extracts were subjected to IP with anti-FASN or anti-TRIM21 antibodies and further analyzed by WB with indicated antibodies. Normal IgG was used as a negative control. Whole-cell lysates were used as a positive control (Input). F The cell extracts from BC cells were subjected to IP with anti-FASN antibody and further analyzed by WB with indicated antibodies. Normal IgG was used as a negative control. Whole-cell lysates were used as a positive control (Input). G The extracts from M231 cells stably expressing FLAG-Vc or FLAG-TC2N (full-length and truncation) were subjected to IP with anti-Flag antibody. Elutes were resolved using SDS-PAGE and silver-stained. Block arrows indicate the bands of TC2N. H The extracts from M231 cells stably expressing FLAG-Vc or FLAG-TC2N (full-length and truncation) were subjected to IP with anti-FASN antibody and further analyzed by WB with indicated antibodies. Normal IgG was used as a negative control. Whole-cell lysates were used as a positive control (Input)