Several endogenous negative regulators of TLR signaling have been described for the MyD88-dependent pathway. MyD88s is the short form of MyD88 and its overexpression inhibits IL-1- and LPS- but not TNF-induced NF-kB activation (Janssens et al. 2003). Another inhibitor of the MyD88-mediated pathway is IRAK-M, a member of the IRAK kinase family (Wesche et al. 1999), which has been shown to block the formation of IRAK1-TRAF6 complexes (Kobayashi et al. 2002). A different level of regulation occurs through SOCS1, one of eight members of the SOCS family important in suppressing cytokine signaling (Alexander 2002). SOCS1 represses LPS-induced NF-kB activation in a TLR4- and MD2-dependent manner (Kinjyo et al. 2002), and Mansell et al. demonstrated recently that SOCS1 is required for the ubiquitin-proteasome-mediated degradation of Mal (Mansell et al. 2006). The inhibitory effect of
SOCS1 on TLR signaling can also be indirect by blocking type I IFN signaling itself (Baetz et al. 2004; Gingras et al. 2004). Several additional negative regulators of the MyD88 pathway have been described, including PI3K (Fukao et al. 2002), Tollip (Zhang and Ghosh 2002), A20 (Boone et al. 2004), ST2 (Brint et al. 2002), SIGIRR (Wald et al. 2003), and RIP105 (Divanovic et al. 2005), all acting at different levels of the intracellular cascade.
Much less is known about negative regulation of the TRIF-IRF3 response. Carty et al. recently demonstrated that the fifth TIR-domain containing adapter SARM acts as a negative regulator of TRIF signaling (Carty et al. 2006). SARM interacts directly with TRIF leading to a block in gene induction downstream of TRIF. SARM does not target the MyD88 pathway. Knockdown of SARM by siRNA leads to enhanced TRIF-dependent cytokine and chemokine induction.
As discussed above, the IKK-related kinases TBK1 and IKKe are involved in IRF3 activation downstream of TRIF. SIKE (for suppressor of IKKe) is a protein that interacts with both TBK1 and IKKe and dissociates from them upon viral infection or TLR3 stimulation (Huang et al. 2005). Overexpression of SIKE blocks the interaction of TBK1 and IKKe with TRIF and IRF3, but does not influence the interaction of TRIF with TRAF6 or Ripl, essential for NF-kB activation. siRNA targeting of SIKE potentiated virus- and TLR3-induced IRF3 responses. Very recently, Saitoh and colleagues demonstrated that the peptidyl prolyl isomerase Pinl also negatively regulates the IRF3 pathway. Pinl associates with activated IRF3 and promotes the ubiquitin-mediated degradation by the proteosome. Phosphorylation of IRF3 on Ser339/Pro440 upon stimulation with poly (I:C), LPS or Newcastle virus is associated with this destabilization of IRF3 (Saitoh et al. 2006). IRF3 and Pinl interact only when IRF3 is phosphory-lated on Ser339. Ectopic expression of Pinl blocks IRF3 activation and IFN-P production downstream of TLR3 and 4 and the RIG-I pathway. As expected, Pinl-deficient mice produce much more IFN-P in response to dsRNA compared to wild type mice in vivo.
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