An Inside Job?

Abstract

Toll-like receptor (TLR) 4 interacts with the TIRAP-MyD88 pair of adaptor proteins to stimulate the production of inflammatory cytokines and with the TRAM-TRIF pair to stimulate the production of type I interferons (see Watts). Curious about how TLR4 coordinates activation of these two signaling pathways, Kagan et al . used tagged constructs to analyze TLR4 location in macrophages and found that it was present both at the plasma membrane and in early endosomes. Inhibition of dynamin-dependent endocytosis prevented lipopolysaccharide (LPS)-induced internalization of endogenous TLR4 and blocked TRAM-TRIF-dependent phosphorylation of the transcription factor IRF3 (and the expression of target genes), whereas TIRAP-MyD88-dependent phosphorylation of p38 mitogen-activated protein kinase and IκBα degradation were unaffected. Whereas fluorescently labeled TIRAP localized to the plasma membrane, fluorescently labeled TRAM was present at both the plasma membrane and in early endosomes. Overall TRAM localization depended on the first 20 residues, a bipartite motif consisting of a previously characterized myristoylation motif (the first 7 residues) followed by a region enriched in basic and aromatic residues, with the myristoylation motif alone sufficient for localization to endosomes. Analyses of LPS-induced cytokine production by TRAM localization mutants indicated that TLR4 signaling through TRAM-TRIF took place in endosomes. Noting that no known TLRs stimulate type I interferon production from the plasma membrane (TLR4 had been thought to be the lone exception), the authors determined that localization of TRAF3 (an adaptor critical for type I interferon production) was solely cytosolic. Moreover, targeting of TRAF3 to the plasma membrane enabled interferon-β induction by TLR2, which isn’t usually involved in this pathway. The authors thus propose that TLR4 stimulates TIRAP-MyD88 signaling from the cell surface and initiates TRAM-TRIF signaling only after internalization. J. C. Kagan, T. Su, T. Horng, A. Chow, S. Akira, R. Medzhitov, TRAM couples endocytosis of Toll-like receptor 4 to the induction of interferon-β. Nat Immunol. 9 , 361-368 (2008). [PubMed] C. Watts, Location, location, location: Identifying the neighborhoods of LPS signaling. Nat Immunol . 9 , 343-345 (2008). [PubMed]