Regulation of TLR4 signaling through the TRAF6/sNASP axis by reversible phosphorylation mediated by CK2 and PP4

Recognition of invading pathogens by Toll-like receptors (TLRs) activates innate immunity through signaling pathways that involved multiple protein kinases and phosphatases. We previously demonstrated that somatic nuclear autoantigenic sperm protein (sNASP) binds to TNF receptor–associated factor 6 (TRAF6) in the resting state. Upon TLR4 activation, a signaling complex consisting of TRAF6, sNASP, interleukin (IL)-1 receptor–associated kinase 4, and casein kinase 2 (CK2) is formed. CK2 then phosphorylates sNASP to release phospho-sNASP (p-sNASP) from TRAF6, initiating downstream signaling pathways. Here, we showed that protein phosphatase 4 (PP4) is the specific sNASP phosphatase that negatively regulates TLR4-induced TRAF6 activation and its downstream signaling pathway. Mechanistically, PP4 is directly recruited by phosphorylated sNASP to dephosphorylate p-sNASP to terminate TRAF6 activation. Ectopic expression of PP4 specifically inhibited sNASP-dependent proinflammatory cytokine production and downstream signaling following bacterial lipopolysaccharide (LPS) treatment, whereas silencing PP4 had the opposite effect. Primary macrophages and mice infected with recombinant adenovirus carrying a gene encoding PP4 (Ad-PP4) showed significant reduction in IL-6 and TNF-α production. Survival of Ad-PP4–infected mice was markedly increased due to a better ability to clear bacteria in a sepsis model. These results indicate that the serine/threonine phosphatase PP4 functions as a negative regulator of innate immunity by regulating the binding of sNASP to TRAF6.

New Role for DCR-1/Dicer in Caenorhabditis elegans Innate Immunity against the Highly Virulent Bacterium Bacillus thuringiensis DB27

ABSTRACTBacillus thuringiensisproduces toxins that target invertebrates, includingCaenorhabditis elegans. Virulence ofBacillusstrains is often highly specific, such thatB. thuringiensisstrain DB27 is highly pathogenic toC. elegansbut shows no virulence for another model nematode,Pristionchus pacificus. To uncover the underlying mechanisms of the differential responses of the two nematodes toB. thuringiensisDB27 and to reveal theC. elegansdefense mechanisms against this pathogen, we conducted a genetic screen forC. elegansmutants resistant toB. thuringiensisDB27. Here, we describe aB. thuringiensisDB27-resistantC. elegansmutant that is identical tonasp-1, which encodes theC. eleganshomolog of the nuclear-autoantigenic-sperm protein. Gene expression analysis indicated a substantial overlap between the genes downregulated in thenasp-1mutant and targets ofC. elegansdcr-1/Dicer, suggesting thatdcr-1is repressed innasp-1mutants, which was confirmed by quantitative PCR. Consistent with this, thenasp-1mutant exhibits RNA interference (RNAi) deficiency and reduced longevity similar to those of adcr-1mutant. Building on these surprising findings, we further explored a potential role fordcr-1inC. elegansinnate immunity. We show thatdcr-1mutant alleles deficient in microRNA (miRNA) processing, but not those deficient only in RNAi, are resistant toB. thuringiensisDB27. Furthermore,dcr-1overexpression rescues thenasp-1mutant's resistance, suggesting that repression ofdcr-1determines thenasp-1mutant's resistance. Additionally, we identified the collagen-encoding genecol-92as one of the downstream effectors ofnasp-1that play an important role in resistance to DB27. Taken together, these results uncover a previously unknown role for DCR-1/Dicer inC. elegansantibacterial immunity that is largely associated with miRNA processing.