Citrullination of NF-κB p65 promotes its nuclear localization and TLR-induced expression of IL-1β and TNFα

Abstract

Citrullination converts peptidyl-arginine to peptidyl-citrulline and is mediated by the enzymes peptidyl arginine deiminases (PADs), including PAD1–4 and PAD6. The physiological role of citrullination in immune cells is poorly understood. Here we report that suppression of PAD activity attenuates TLR-induced expression of IL-1β and TNFα by neutrophils in vivo and in vitro but not their global transcription activity. Mechanistically, PAD4 directly citrullinates NF-kB p65 and enhances the interaction of p65 with importin α3, which brings p65 into the nucleus. The citrullination-enhanced interaction of p65 with importin α3, and its nuclear translocation and transcriptional activity can be attributed to citrullination of four arginine residues located in the Rel homology domain of p65. Furthermore, a rheumatoid arthritis-prone variant of PAD4, carrying three missense mutations, is more efficient in interacting with p65 and enhancing NF-kB activity. Together, these data demonstrate a critical role of citrullination in NF-kB-dependent expression of IL-1β and TNFα.

Frontline Science: Anthrax lethal toxin-induced, NLRP1-mediated IL-1β release is a neutrophil and PAD4-dependent event

“…PAD4 impacts NLRP3 inflammasome function, 71 and could thus also possibly alter NLRP1 responses. Furthermore, citrullination of NF‐κB has recently been shown to control TLR‐mediated expression of cytokines, suggesting a direct role for PAD4 activity in upregulation of cytokines by a variety of signals, possibly including signal 1 in our studies 72 . The fact that we have not found neutrophils to produce either IL‐1β or form NETs in response to LT in vitro complicates the hypothesis that PAD4‐mediated NET formation plays a role in the IL‐1β response, although the heterogeneity of neutrophils in both blood and marrow cannot be underestimated 73 .…”

Peptidylarginine deiminases 2 and 4 modulate innate and adaptive immune responses in TLR-7–dependent lupus

“…An imbalance of NET formation and degradation occurs in SLE, and this phenomenon may play a role in augmenting the half-life of externalized modified autoantigens that, in a predisposed host, would contribute to fueling the immune system, which would promote type I IFN responses, inflammasome activation, aberrant adaptive immunity, and tissue damage, including vascular injury (20,21,(23)(24)(25). In addition to the role in NET formation, PAD4 enhances the activity of NF-κB in neutrophils, independently of changes in histone citrullination, with potential therapeutic implications for TLR-mediated inflammation (26).…”

Recombinant human ADAMTS13 treatment and anti-NET strategies enhance skin allograft survival in mice

“…To the general field of organ transplant, inhibiting NET generation, rather than cleaving NETs, may represent an unequivocal beneficial approach to enhance allograft survival and tolerance. Indeed, apart from chromatin decondensation for NETosis, PAD4 also augments the expression of proinflammatory cytokines, IL‐1β and tumor necrosis factor‐α, by citrullinating NF‐κB p65 subunit, thereby promoting its nuclear translocation in neutrophils . In macrophages, PAD2 and PAD4 are required for the assembly of NLRP3 inflammasome and IL‐1β release .…”

“…Indeed, apart from chromatin decondensation for NETosis, PAD4 also augments the expression of proinflammatory cytokines, IL-1β and tumor necrosis factor-α, by citrullinating NF-κB p65 subunit, thereby promoting its nuclear translocation in neutrophils. 35 In macrophages, PAD2 and PAD4 are required for the assembly of NLRP3 inflammasome and IL-1β release. 36 Interestingly, recent results indicate that NLRP3 inflammasome in neutrophils is important in NET formation, as well.…”