Coordinate regulation of TPL-2 and NF-κB signaling in macrophages by NF-κB1 p105

Abstract

The role of IκB kinase (IKK)-induced proteolysis of NF-κB1 p105 in innate immune signaling was investigated using macrophages from Nfkb1SSAA/SSAA mice, in which the IKK-target serines on p105 are mutated to alanines. We found that the IKK/p105 signaling pathway was essential for TPL-2 kinase activation of ERK MAP kinase, and modulated the activation of NF-κB. Nfkb1SSAA mutation prevented the agonist-induced release of TPL-2 from its inhibitor p105, which blocked activation of ERK by lipopolysaccharide (LPS), TNF, CpG, Pam3CSK, poly-I/C, flagellin and R848. Nfkb1SSAA mutation also prevented LPS-induced processing of p105 to p50, and reduced p50 levels, in addition to decreasing the nuclear translocation of RelA and cRel. Reduced p50 in Nfkb1SSAA/SSAA macrophages significantly decreased LPS-induction of the IκBζ-regulated Il6 and Csf2 genes. LPS upregulation of Il12a and Il12b mRNAs was also impaired, although specific blockade of TPL-2 signaling increased expression of these genes at late time points. Activation of TPL-2/ERK signaling by IKK-induced p105 proteolysis, therefore, induced a negative feedback loop to down-regulate NF-κB-dependent expression of the pro-inflammatory cytokine IL-12. Unexpectedly, TPL-2 promoted soluble TNF production independently of IKK-induced p105 phosphorylation and its ability to activate ERK, which has important implications for the development of anti-inflammatory drugs targeting TPL-2.