TLR8-mediated NF-κB and JNK activation are TAK1-independent and MEKK3-dependent

Abstract

TLR8-mediated NF-κB and IRF7 activation are abolished in human IRAK-deficient 293 cells and IRAK4-deficient fibroblast cells. Both wild-type and kinase-inactive mutants of IRAK and IRAK4, respectively, restored TLR8-mediated NF-κB and IRF7 activation in the IRAK- and IRAK4-deficient cells, indicating that the kinase activity of IRAK and IRAK4 is probably redundant for TLR8-mediated signaling. We recently found that TLR8 mediates a unique NF-κB activation pathway in human 293 cells and mouse embryonic fibroblasts, accompanied only by IκBα phosphorylation and not IκBα degradation, whereas interleukin (IL)-1 stimulation causes both IκBα phosphorylation and degradation. The intermediate signaling events mediated by IL-1 (including IRAK modifications and degradation and TAK1 activation) were not detected in cells stimulated by TLR8 ligands. TLR8 ligands trigger similar levels of IκBα phosphorylation and NF-κB and JNK activation in TAK1-/- mouse embryo fibroblasts (MEFs) as compared with wild-type MEFs, whereas lack of TAK1 results in reduced IL-1-mediated NF-κB activation and abolished IL-1-induced JNK activation. The above results indicate that although TLR8-mediated NF-κB and JNK activation are IRAK-dependent, they do not require IRAK modification and are TAK1-independent. On the other hand, TLR8-mediated IκBα phosphorylation, NF-κB, and JNK activation are completely abolished in MEKK3-/- MEFs, whereas IL-1-mediated signaling was only moderately reduced in these deficient MEFs as compared with wild-type cells. The differences between IL-1R- and TLR8-mediated NF-κB activation are also reflected at the level of IκB kinase (IKK) complex. TLR8 ligands induced IKKγ phosphorylation, whereas IKKα/β phosphorylation and IKKγ ubiquitination that can be induced by IL-1 were not detected in cells treated with TLR8 ligands. We postulate that TLR8-mediated MEKK3-dependent IKKγ phosphorylation might play an important role in the activation of IKK complex, leading to IκBα phosphorylation. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.