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. 2003 Oct 6;198(7):1043–1055. doi: 10.1084/jem.20031023

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Copyright © 2003, The Rockefeller University Press

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Figure 3. — TRAM interacts with IRF-3 and CBP and signals via IKKɛ and TBK1. (a) IRF-3–GFP–expressing HEK293 cells were plated on 35-mm glass-bottom sterile tissue culture dishes and transiently transfected with 1 μg of Flag-tagged TRAM, TRIF, or pCDNA3.1 and visualized 24 h later by confocal microscopy. (b) 293T cells were transfected with 4 μg of Flag-TRAM with or without a plasmid encoding IRF-3 (untagged) as indicated. 24 h later, whole cell lysates were immunoprecipitated with anti–IRF-3, anti-Flag, or anti-CBP, and the immunoprecipitated complexes were immunoblotted for Flag-tagged TRAM and IRF-3. Whole cell lysates (WCL) were also analyzed for Flag-tagged proteins. (c) HEK293 cells were transfected with the RANTES luciferase reporter gene and TRAM (20 ng) and cotransfected with increasing concentrations of IKKɛ-k38a, TBK1-k38a, or IRF3-ΔN from 10, 20, 30, 40, 60, and 80 ng. Luciferase reporter gene activity was measured 24 h after transfection.