Toll-like receptors (TLRs) are a major family of pattern recognition receptors, and they play a crucial role in innate immune responses. Activation of TLR4 signaling at the plasma membrane by its ligand lipopolysaccharide (LPS) stimulates a proinflammatory pathway dependent on the E3 ubiquitin ligase TRAF6 (tumor necrosis factor receptor-associated factor 6) and the kinase TAK1 (transforming growth factor β-activated kinase 1), whereas TLR4 signaling at endosomes stimulates the production of type I interferons (IFNs) through a pathway that depends on TRAF3 and the kinase TBK1 (TANK-binding kinase-1). We found that the nonreceptor tyrosine kinase Syk partially mediated the endocytosis of TLR4, but it also played a dual role in TLR4-mediated signaling. LPS-dependent stimulation of TLR4 in Syk-deficient macrophages led to enhanced activation of TAK1 and increased production of proinflammatory cytokines compared to that in wild-type macrophages. In contrast, Syk-deficient macrophages exhibited decreased TLR4-dependent activation of TBK1 signaling and production of type I IFNs. We found that Syk was present in both TRAF6-and TRAF3-containing signaling complexes; however, the LPS-dependent, lysine 63-linked ubiquitination of TRAF6 and TRAF3 was oppositely regulated by Syk. We identified the domains of Syk that interacted with TRAF3, TRAF6, TAK1, and TBK1, factors activated by multiple TLRs, which suggests that Syk may act as a common regulator of various TLR responses. Together, our results demonstrate the opposing regulatory roles of Syk in TLR-mediated TRAF6 and TRAF3 signaling pathways, which suggests that Syk may fine-tune the innate immune response to lessen inflammation.
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