Negative regulation of NF-κB p65 activity by serine 536 phosphorylation

Abstract

Nuclear factor κB (NF-κB) is a master regulator of inflammation and cell death. Whereas most of the activity of NF-κB is regulated through the inhibitor of kB (IκB) kinase (IKK)–dependent degradation of IκB, IKK also phosphorylates subunits of NF-κB. Here, we investigated the contribution of the phosphorylation of the NF-κ B subunit p65 at the IKK phosphorylation site serine 536 (Ser536) in humans, which is thought to be required for the activation and nuclear translocation of NF-κB. Through experiments with knock-in mice (S534A mice) expressing a mutant p65 with an alanine-to-serine substitution at position 534 (the murine homolog of human Ser536), we observed increased expression of NF-κ B–dependent genes after injection of mice with the inflammatory stimulus lipopolysaccharide (LPS) or exposure to gamma irradiation, and the enhanced gene expression was most pronounced at late time points. Compared to wild-type mice, S534A mice displayed increased mortality after injection with LPS. Increased NF-κ B signaling in the S534A mice was at least in part explained by the increased stability of the S534A p65 protein compared to that of the Ser534-phosphorylated wild-type protein. Together, our results suggest that Ser534 phosphorylation of p65 in mice (and, by extension, Ser536 phosphorylation of human p65) is not required for its nuclear translocation, but instead inhibits NF-κ B signaling to prevent deleterious inflammation.

CDK7 inhibition suppresses rheumatoid arthritis inflammation via blockage of NF‐κB activation and IL‐1β/IL‐6 secretion

“…In our study, we showed that CDK7 inhibition suppressed NF‐κB signalling pathway activation and p65 nuclear translocation in RA FLS (Figs and ), indicating that reduction of IL‐1β/IL‐6 secretion by CDK7 inhibition might be due to NF‐κB signalling pathway blockage. Furthermore, besides phosphorylation and subsequent degradation of inhibitory molecular, protein kinases are also required for optimal NF‐κB activation by targeting functional domains of NF‐κB protein themselves . Our data also displayed that CDK7 inhibitor BS‐181 greatly decreased the phosphorylation of p65 (Fig.…”

Epstein‐Barr virus tegument protein BGLF2 inhibits NF‐κB activity by preventing p65 Ser536 phosphorylation

“…kinase-1, and the interaction between NF-kB p65 and mitogen-and stress-activated protein kinase-1 results in fully functional NF-kB, which can recruit transcriptional coactivators, such as p300 and CREB-binding protein, to drive transcription of NF-kB-regulated genes (61)(62)(63). To further investigate whether BGLF2 affects the phosphorylation of p65, HEK293T cells were transfected with either plasmid pBGLF2-Flag or vector control pFlag-N1.…”

Disease-modifying effects of orally bioavailable NF-κB inhibitors in dystrophin-deficient muscle

“…An IκB-independent, alternative NF-κB pathway (altNF-κB) exists that involves the activation of RelB-p52 heterodimers by IKKα-induced proteolytic cleavage of p100 into p52 (3). Additionally, phosphorylation of a pool of IκB-independent p65 on Ser536 has been reported to result in p65-p65 homodimer formation and activation of genes distinct from cNF-κB activation (6); however, recent evidence suggests this modification serves as a brake on p65-dependent transcription (7).…”