Evidence for the ancient origin of the NF-κB/IκB cascade: Its archaic role in pathogen infection and immunity

The evolutionary conservation of the NF-κB transcription factors, from Drosophila to humans, underscores its pivotal role in immune response. Unexpectedly, the canonical NF-κB signaling pathway is not functional in the immune system of Caenorhabditis elegans. Therefore, the ancient origin of the NF-κB signaling pathway is still unknown. Here, we report the discovery and characterization of a primitive and functional NF-κB/IκB pathway in the immune defense of a "living fossil," the horseshoe crab, Carcinoscorpius rotundicauda. The ancient NF-κB/IκB homologues, CrNFκB, CrRelish, and CrIκB, share numerous signature motifs with their vertebrate orthologues. CrNFκB recognizes both horseshoe crab and mammalian κB response elements. CrIκB interacts with CrNFκB and inhibits its nuclear translocation and DNA-binding activity. The activation of the CrNFκB is autoregulated by a feedback mechanism mediated by CrIκB, the natural inhibitor of CrNFκB. We further show that Gram-negative bacteria infection causes rapid degradation of CrIκB and nuclear translocation of CrNFκB. Infection also leads to an increase in the κB-binding activity and up-regulation of immune-related gene expression, like inducible nitric oxide synthase and Factor C, an LPS-activated serine protease. Altogether, our study shows that, although absent in C. elegans, the NF-κB/IκB signaling cascade remains well conserved from horseshoe crab to humans, playing an archaic but fundamental role in regulating the expression of critical immune defense molecules.