UbcH5 is required for IKK activation by TNFα, but not IL-1β
(A) Tetracycline (Tet) was used to induce the expression of shRNA against both UbcH5b and UbcH5c. After stimulation with GST-TNFα or IL-1β for the indicated time, the activation of IKK was analyzed by immunoblotting with an IκBα antibody or by IKK activity assay. (B) U2OS cells expressing UbcH5 shRNA and RNAi-resistant Flag-UbcH5c (left) or Flag-UbcH5c (C85A; right) were treated with tetracycline for 7 days. The cells were stimulated with GST-TNFα for the indicated time and then IKK activation was measured. IKK activity was quantified using PhosphoImager and the fold activation as compared to unstimulated cells is shown.