Fig. 3.

OTUD1 interacts with MCL1 and promotes MCL1 deubiquitination. a The indicated plasmids were transfected into 293T cells. Next, the cell lysates were immunoprecipitated with an anti-Flag antibody. The lysates (TCL) and the immunoprecipitates were immunoblotted with the indicated antibodies. b The indicated plasmids were transfected into 293T cells. The cell lysates were then immunoprecipitated with an anti-HA antibody. The lysates (TCL) and the immunoprecipitates were immunoblotted with indicated antibodies. c The empty vector, OTUD1 or its enzymatic domain (amino acids 309-481) were co-transfected with MCL1-HA into 293T cells. The cell lysates were immunoprecipitated with an anti-Flag antibody. The lysates (TCL) and the immunoprecipitates were immunoblotted with the indicated antibodies. d The empty vector, OTUD1 or its enzymatic activity mutant C320S-expressing plasmid was co-transfected with MCL1-HA and Flag-ubiquitin expression plasmids into 293T cells. The cells were treated with MG132 (10 µM) for 6 h, and then the cells were lysed in RIPA buffer and the cell lysates were immunoprecipitated with anti-HA antibody. The lysates and the immunoprecipitates were subjected to SDS-PAGE and analysed by immunoblotting with the indicated antibodies