Fig. 5: XIAP is a novel ubiquitin E3 ligase for YTHDC1.
From: XIAP promotes metastasis of bladder cancer cells by ubiquitylating YTHDC1
HA-XIAP and Flag-YTHDC1 were co-transfected into UMUC3 cells. Immunoprecipitation was performed using Anti-HA-tag mAb-Magnetic Beads (A) or Anti-DDDDK-tag mAb-Magnetic Beads (B) to detect the interaction between XIAP and YTHDC1. C UMUC3 cell lysates were subject to immunoprecipitation with control IgG and anti-XIAP antibodies. D HA-XIAP and Flag-YTHDC1 were co-transfected and their co-localization was observed by confocal microscopy. E, F Schematic illustration of YTHDC1 structure (E). UMUC3 cells were transfected with the indicated constructs and co-immunoprecipitated with Anti-DDDDK-tag mAb-Magnetic Beads (F). G YTHDC1 mutant plasmids were constructed at amino acids 372, 469 and 472. H The plasmids HA-XIAP, Myc-ub and Flag-YTHDC1 (Vec, WT, K372R, K469R, K472R, 3KR) were transfected into UMUC3 cells and the level of ubiquitination was detected by immunoprecipitation. I The plasmids Flag-YTHDC1, Flag-YTHDC1-3KR and HA-XIAP were transfected into the cells, and YTHDC1 degradation rate was detected by western blotting.
