TLR3 signaling complex forms by cooperative receptor dimerization

Toll-like receptors (TLRs) initiate immune responses by recognizing pathogen-associated molecules, but the molecular basis for recognition is poorly understood. In particular, it is unclear how receptor-ligand interactions lead to the initiation of downstream signaling. Here, we describe the mechani...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2008-01, Vol.105 (1), p.258-263
Main Authors: Leonard, Joshua N, Ghirlando, Rodolfo, Askins, Janine, Bell, Jessica K, Margulies, David H, Davies, David R, Segal, David M
Format: Article
Language:English
Subjects:
Binding Sites
Biological Sciences
Buffers
Cell Line
Cell lines
Cell Separation
Chromatography, Gel
Cross-Linking Reagents - pharmacology
Dimerization
Dimers
DNA, Single-Stranded - chemistry
Double stranded RNA
Endosomes
Flow Cytometry
HEK293 cells
Humans
Hydrogen-Ion Concentration
Immune system
Immunology
Ligands
Models, Biological
Molecules
Oligonucleotides
Oligonucleotides - chemistry
Receptors
Ribonucleic acid
RNA
RNA, Double-Stranded - chemistry
Studies
Toll like receptors
Toll-Like Receptor 3 - chemistry
Toll-Like Receptor 3 - metabolism
Toll-Like Receptor 4 - chemistry
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Summary:Toll-like receptors (TLRs) initiate immune responses by recognizing pathogen-associated molecules, but the molecular basis for recognition is poorly understood. In particular, it is unclear how receptor-ligand interactions lead to the initiation of downstream signaling. Here, we describe the mechanism by which TLR3 recognizes its ligand, double-stranded RNA (dsRNA), and forms an active signaling complex. We show that dsRNA binds saturably, specifically, and reversibly to a defined ligand-binding site (or sites) on the TLR3 ectodomain (TLR3ecd). Binding affinities increase with both buffer acidity and ligand size. Purified TLR3ecd protein is exclusively monomeric in solution, but through a highly cooperative process, it forms dimers when bound to dsRNA, and multiple TLR3ecd dimers bind to long dsRNA strands. The smallest dsRNA oligonucleotides that form stable complexes with TLR3ecd (40-50 bp) each bind one TLR3ecd dimer, and these are also the smallest oligonucleotides that efficiently activate TLR3 in cells. We conclude that TLR3 assembles on dsRNA as stable dimers and that the minimal signaling unit is one TLR3 dimer.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.0710779105