Coexpression of vGPCR with dominant negative Rac constructs using an IRES system. (A) Schematic representation of the bicistronic constructs encoding the vGPCR and GFP, RacN17, or PAKN. (B) Immunofluorescence demonstrating coexpression both of vGPCR and GFP, RacN17, or PAKN in COS-7 cells transfected with IRES constructs. (C) COS-7 cells coexpressing both vGPCR and RacN17 or PAKN fail to stimulate NF-κB or induce IL-6 transcription. 293T cells coexpressing both vGPCR and RacN17 or PAKN fail to induce IL-6 secretion. Data represent the mean ± SEM of triplicate samples from a typical experiment and are expressed as percent of luciferase activity, CAT activity, or IL-6 secretion levels with respect to vGPCR-I-GFP–expressing cells.