Fig. 5.

Repression of IKK activity in intact cells by endogenous NOS activity. Jurkat T cells were treated with 1 mM l-NMMA for 4 h followed by stimulation with 10 ng/ml TNFα for 10 min or mock manipulations. Selected dishes were treated with TNFα alone. (A) IKK activity was assessed in an in vitro kinase assay. IB, anti-IKKβ immunoblot. (B) Lysates were subjected to biotin derivatization, and biotinylated IKKβ was detected after immunoprecipitation of IKKβ and Western blotting by using streptavidin–horseradish peroxidase. In control samples, reduction by ascorbate (–vit C) or labeling with N-(3-malemidylpropionyl)biocytin (–biotin) was omitted. IB, anti-IKKβ immunoblot.