Two tyrosine residues of toll-like receptor 3 trigger different steps of NF-κB activation

Innate immune response to viral infection is often triggered by Toll-like receptor 3 (TLR3)-mediated signaling by double-stranded (ds) RNA, which culminates in the activation of the transcription factor NF-κB and induction of NF-κB-driven genes. We demonstrated that dsRNA-induced phosphorylation of two specific tyrosine residues, 759 and 858, of TLR3 was necessary and sufficient for complete activation of the NF-κB pathway. When Tyr-759 of TLR3 was mutated, gene induction was inhibited, although NF-κB was partially activated. It was released from IκB and translocated to the nucleus but failed to bind to the κB site of the target A20 gene promoter. This defect could be attributed to incomplete phosphorylation of the RelA (p65) subunit of NF-κB, as revealed by two-dimensional gel analyses of p65, isolated from dsRNA-treated cells expressing either wild type TLR3 or the Tyr-759 → Phe mutant TLR3. Thus, two phosphotyrosine residues of TLR3 activate two distinct pathways, one leading to NF-κB release and the other leading to its phosphorylation.