NF-κB RelA Phosphorylation Regulates RelA Acetylation

Abstract

The NF-B/Rel family of transcription factors plays a key role in regulating inflammatory and immune responses and other programs of cell growth and survival. The five known mammalian Rel genes encode seven Rel-related proteins: RelA/p65; p105 and its processing product, p50; p100 and its processing product, p52; c-Rel; and RelB. Each contains an N-terminal Rel homology domain (ϳ300 amino acids) that mediates DNA binding, dimerization, and interaction with the IB family of NF-B/Rel inhibitors. RelA, c-RelA, and RelB contain C-terminal transactivation domains, but p50 and p52 do not. Each NF-B/Rel protein forms different homo-or heterodimers with other members of the family, which may contribute to the activation of specific target genes (1, 5).The prototypical NF-B complex is a p50/RelA heterodimer. NF-B is largely sequestered in the cytoplasm through its association with an IB inhibitor. Nuclear NF-B expression is induced by various stimuli, including proinflammatory cytokines, growth factors, DNA-damaging agents, and viral proteins (13). The activation of NF-B can be divided into two phases. The first phase involves cytoplasmic events culminating in the activation of the IB kinases (IKK1 and IKK2). These kinases promote N-terminal phosphorylation of serines 32 and 36 in IB␣, leading to its polyubiquitylation and proteasome-mediated degradation. The liberated NF-B complex rapidly translocates to the nucleus, ending the first phase (13). The second phase occurs primarily in the nucleus and involves posttranslational modification of the NF-B transcription factor complex or relevant histones surrounding NF-B target genes (5). These modifications determine both the strength and duration of the NF-B-mediated transcriptional response (5).One of the nuclear events is the reversible acetylation of RelA (4). Endogenous RelA is acetylated in a stimulus-coupled manner after activation of cells with tumor necrosis factor alpha (TNF-␣), phorbol myristate acetate, or other stimuli at multiple sites, including lysines 122, 123, 218, 221, and 310 (4, 17). The acetyltransferases p300 and CBP appear to play a major role in the in vivo acetylation of RelA (6,17). Sitespecific acetylation of RelA regulates discrete biological actions of the NF-B complex (5, 6). For example, acetylation of lysine 221 by p300/CBP increases the DNA binding affinity of RelA for the B enhancer and, together with acetylation of lysine 218, impairs assembly of RelA with newly synthesized IB␣, which shuttles in and out of the nucleus. Acetylation of lysine 310 does not modulate DNA binding or IB␣ assembly but markedly enhances the transcriptional activity of NF-B. Deacetylation of lysine 310 by histone deacetylase 3 (HDAC3) or SIRT1 inhibits the transcriptional activity of RelA and augments cellular apoptosis in response to 32). While it is clear that signal-coupled acetylation of RelA participates in the nuclear regulation of NF-B action (4, 17), many unanswered questions remain. Chief among these is how the acetylation of RelA is regulated.

Gallic Acid Suppresses Lipopolysaccharide-Induced Nuclear Factor-κB Signaling by Preventing RelA Acetylation in A549 Lung Cancer Cells

“…Thus, our study provides firm evidence to support the idea that gallic acid suppresses p300/ CBP-mediated p65 acetylation. The acetylation of p65 is known to be enhanced by prior phosphorylation of p65 at serine 276 or serine 536 (26). By immunoprecipitation and Western blot analysis, we observed that gallic acid treatment leads to the inhibition of p65 phosphorylation (data not shown).…”

“…We conclude that gallic acid prevents the hyperacetylation of p65 by inhibiting the HAT activity of p300/CBP. p300-Mediated RelA Acetylation Is Critical for NF-κB Function Because p300-mediated RelA acetylation is critical for the maintenance of NF-κB function (12,26), we first examined the effect of gallic acid on the LPS-induced DNA binding activity of NF-κB by performing electrophoretic mobility shift assays (EMSA). As shown in Fig.…”

Inhibition of histone deacetylases prevents cytokine-induced toxicity in beta cells

“…Stimulus-induced phosphorylation of p65 at serine residue 276 determines whether it associates with co-activators or co-repressors, as phosphorylation facilitates CREB binding protein/p300 association, while affinity of HDAC-1 for p65 is decreased [44]. This phosphorylation precedes NFκB acetylation, leading to increased expression of NFκB-responsive genes [45]. We have previously shown that serine 276 does indeed play an important role in cytokine-stimulated NFκB-mediated gene expression in beta cells [25] setting the stage for cytokine-induced NFκB acetylation.…”

Copine-I represses NF-κB transcription by endoproteolysis of p65

“…To effectively recruit co-activator complexes, the p65 subunit of NF-kB must be posttranslationally modified on specific residues. Phosphorylation of p65 at serine 276 or 536 is necessary for exchanging co-repressor complexes for co-activator complexes (Zhong et al, 2002;Chen et al, 2005;Hoberg et al, 2006). Our laboratory has shown that the activation and repression of NF-kB-responsive genes is a dynamic process, with continual exchanges between co-activator and co-repressor complexes (Hoberg et al, , 2006.…”