Regulation of NF-κB signaling by ASK1 through interaction with NIK

(A) Overexpressed NIK and ASK1proteins bind directly. HA-tagged ASK1 and FLAG-NIK were tran-siently co-transfected into BOSC 23 cells, and the lysates were immuno-precipitated with appropriate antibodies. Samples were subjected to Western blotting analysis using appropriate antibodies. The expressionof transfected plasmids were indicated in the same lysates. IP, immuno-precipitation; IB, immunoblot. (B) NIK binds to the C-terminal domai nof ASK1. BOSC 23 cells were transiently co-transfected with FLAG-tagged NIK and HA-tagged ASK1 wild-type or mutant expression plasmids. Transfected cells were extracted and immunoprecipitated with an anti-HA antibody. The interaction was detected by Western blotting with an anti-FLAG antibody.