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TLR signal via Toll-IL-1R (TIR) homology domain-containing adaptor proteins. One of these adaptors, Toll-IL-1R domain-containing adaptor inducing IFN-beta-related adaptor molecule (TRAM), has been shown to be essential for TLR4 signaling in TRAM(-/-) mice and cell lines. Previously, we showed that MyD88 or Mal dominant-negative constructs did not inhibit LPS induction of cytokines in primary human M-CSF-derived macrophages. A possible explanation was redundancy of the adaptors during LPS signaling. TRAM is a suitable candidate to compensate for these adaptors. To investigate a potential role for TRAM in LPS signaling in human M-CSF-derived macrophages, we engineered an adenoviral construct expressing dominant-negative TRAM-C117H (AdTRAMdn). Synovial fibroblasts (SF) and human umbilical endothelial cells (HUVECs) were used as a nonmyeloid comparison. AdTRAMdn inhibited LPS-induced signaling in SFs and HUVECs, reducing NF-kappaB activation and cytokine production, but did not inhibit LPS signaling in M-CSF-derived human macrophages. Further investigation of other TLR ligands showed that AdTRAMdn was also able to inhibit signaling initiated by lipoteichoic acid, a TLR2 ligand, in SFs and HUVECs and lipoteichoic acid and macrophage-activating lipopeptide 2 signaling was also inhibited in TRAM(-/-) murine embryonic fibroblasts. We conclude that TRAM is an adaptor protein for both TLR4 and TLR2/6 signaling in SFs, HUVECs, and murine embryonic fibroblasts, but cannot demonstrate a role in human macrophages.

Type

Journal article

Journal

J Immunol

Publication Date

15/02/2007

Volume

178

Pages

2148 - 2154

Keywords

Adaptor Proteins, Signal Transducing, Adaptor Proteins, Vesicular Transport, Animals, Cells, Cultured, Embryo, Mammalian, Endothelial Cells, Fibroblasts, Humans, Lipopolysaccharides, Macrophages, Mice, Mice, Knockout, NF-kappa B, Organ Specificity, Receptors, Interleukin, Signal Transduction, Species Specificity, Synovial Membrane, Teichoic Acids, Toll-Like Receptor 2, Toll-Like Receptor 4, Toll-Like Receptor 6, Umbilical Cord