Figure 1.

TRAF2 is required for Smac mimetic (SM)-induced cIAP1 degradation. (a and e) Biotinylated SM was used to purify SM-binding proteins from lysates of MDA-MB-231 (a) and MEFs (e). The presence of co-purified proteins was established by immunoblotting the eluate with the indicated antibodies. (b–d and g) WT and TRAF-knockout MEFs were treated with 100 nM Comp. A, 100 nM Comp. C and 1 μM LBW242 for the indicated time points (min). The presence of the indicated proteins was established by immunoblotting the lysates with the indicated antibodies. Asterisks indicate nonspecific bands. (f) Secreted luciferase reporter assays using TRAF2^{−/−,i−IκB−SR} MEFs. Cells were left untreated or treated with 100 ng/ml doxocycline (Dox) to induce IκB-SR. The medium was analysed for luciferase activity 24 h later. Luciferase activity is shown relative to the uninduced condition. The error bar indicates S.D. of triplicate experiments. (g) TRAF2^{−/−,i−IκB−SR} MEFs were left uninduced or induced with Dox and treated with 1 μM LBW242 for the indicated time points