Toll-like receptor 3 (TLR3) recognizes double-stranded RNA (dsRNA), a molecular signature of most viruses, and triggers inflammatory responses that prevent viral spread. TLR3 ectodomains (ECDs) dimerize on oligonucleotides of at least 40 to 50 base pairs in length, the minimal length required for signal transduction. To establish the molecular basis for ligand binding and signaling, we determined the crystal structure of a complex between two mouse TLR3-ECDs and dsRNA at 3.4 angstrom resolution. Each TLR3-ECD binds dsRNA at two sites located at opposite ends of the TLR3 horseshoe, and an intermolecular contact between the two TLR3-ECD C-terminal domains coordinates and stabilizes the dimer. This juxtaposition could mediate downstream signaling by dimerizing the cytoplasmic Toll interleukin-1 receptor (TIR) domains. The overall shape of the TLR3-ECD does not change upon binding to dsRNA.

The membrane-bound Toll-like receptors (TLRs) trigger innate immune responses following recognition of a wide variety of pathogen-derived compounds. Despite the wide range of ligands recognized by TLRs, the receptors share a common structural framework in their extracellular, ligand-binding domains. These domains all adopt horseshoe-shaped structures built from leucine-rich repeat motifs. Typically, upon ligand binding, two extracellular domains form an “m”-shaped dimer sandwiching the ligand molecule bringing the transmembrane and cytoplasmic domains in close proximity and triggering a downstream signalling cascade. Although the ligand-induced dimerization of these receptors has many common features, the nature of the interactions of the TLR extracellular domains with their ligands varies markedly between TLR paralogs.