Activation of Nuclear Transcription Factor NF-κB by Interleukin-1 Is Accompanied by Casein Kinase II-mediated Phosphorylation of the p65 Subunit

Abstract

In fibroblasts and hepatoma cells, interleukin-1 (IL-1) treatment results in the rapid nuclear accumulation of the transcription factor NF-B, present largely as p65 (RelA)/p50 heterodimers. It is well established that this process is dependent in large part upon the phosphorylation and subsequent degradation of the cytosolic inhibitor IB. We looked for other IL-1-induced modifications of NF-B components and found that, in both cell types, IL-1 stimulation led, within minutes, to phosphorylation of both NF-B p65 and p50. Phosphorylation of p65 was sustained for at least 30 min after addition of the cytokine and occurred principally upon serine residues. Immunoprecipitates of NF-B complexes contained an associated protein kinase, the biochemical characteristics of which were indistinguishable from casein kinase II (CKII). Purified CKII efficiently phosphorylated p65 in vitro, apparently on the same major sites that became phosphorylated in intact IL-1-treated cells. Although IL-1 treatment caused little apparent stimulation of total cellular CKII activity, the fraction that was specifically associated with NF-B complexes was markedly elevated by the cytokine. The association of CKII with NF-B occurred in the cytoplasm, suggesting that this phosphorylation might be involved either in control of translocation of the activated complex or in modulation of its DNA binding properties. Interleukin-1 (IL-1)1 receptors are widely distributed on cells of many lineages (1). Thus IL-1 can exert effects on connective tissues, for example inducing mitogenesis through secretion of PDGF; effects on lymphoid cells, such as inducing up-regulation of surface immunoglobulin and secretion of other cytokines; and effects on hepatocytes, for example inducing secretion of acute phase proteins such as C-reactive protein. All of these responses are triggered through a single type of receptor (IL-1R1) (2), which is the archetype for a diverse family of proteins. Included in this group are the insect Toll proteins (3), which function in the establishment of dorsal-ventral polarity during embryogenesis, and the product of the tobacco N-gene (4), which mediates resistance to the pathogen tobacco mosaic virus. Many IL-1-responsive genes are regulated through activation of Rel family transcription factors, in particular NF-B p65/p50 heterodimers, and indeed, ligation of Toll ultimately signals the activation of Dorsal, a Rel-related protein (5).NF-B is a term used to refer to a group of transcription factors/DNA binding activities that recognize a consensus motif, 5Ј-GGGRNNYYCC-3Ј. All of these factors are dimers composed of subunits that share a common domain prototypically found in c-Rel (6). The transactivating activity of the NF-B factors is regulated by a distinct set of proteins termed IBs, which are characterized by a series of ankyrin repeats. IBs function by sequestering NF-B dimers in the cytosol. (7). A variety of stimuli can induce activation of NF-B (8), including viruses, lipopolysaccharides, and pro-inflammatory cytokines such as ...

Breakpoint Cluster Region–Mediated Inflammation Is Dependent on Casein Kinase II

“…Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis showed that CK2a is a candidate (Table I). Consistently, it was previously shown that CK2a associates with the BCR moiety of BCR-Abl (24,25), and CK2 phosphorylates p65 (10)(11)(12). Therefore, we hypothesized that CK2a associated with not only BCR-Abl but also BCR and that CK2a is a critical kinase for BCR-mediated inflammation development in vivo and in vitro.…”

“…CK2 forms a heterocomplex composed of two catalytic a subunits and two regulatory subunits (9). It was reported that CK2 phosphorylates p65 (10)(11)(12), an action critical for NF-kB-mediated transcription (13,14), although a detailed molecular mechanism of how CK2 regulates the transcription has not been shown.…”

Nuclear Accumulation of cRel following C-Terminal phosphorylation by TBK1/IKKε

“…21). Thus far, inducible phosphorylation of RelA/p65 at serine residues 276 or 311 in the Rel homology domain or serine residues 468, 529, and 536 in the TD have been observed following exposure of cells to various NF-B-inducing agents, culminating in the modulation of nuclear localization, DNA-binding affinity, coactivator/corepressor association, and transactivation capacity (14,(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33). Inducible phosphorylation of cRel was also detected in response to PMA, CD28, or TNF-␣ treatment of T cells (34 -36); phosphorylation of Ser 471 in the TD has been reported to be involved in TNF-␣-induced activation of cRel (34,35), whereas Ser 451 in the TD of murine cRel serves as a phospho-acceptor site, although the physiological function of this event remains to be elucidated (37).…”

La protéine kinase CK2, une enzyme qui cultive la différence

“…partie des kinases qui règlent la dégradation de IκB [26] et stimulent l'activité transcriptionnelle de NF-κB [27,28]. Le fait que le promoteur du gène de CK2α soit régulé par NF-κB permet d'envisager l'existence d'une boucle de rétrocontrôle positif conduisant à une surexpression concertée de CK2α et de NF-κB dans certains cancers [29].…”