Pim-1 controls NF-κB signalling by stabilizing RelA/p65

Abstract

Post-translational modification and degradation of proteins by the ubiquitin-proteasome system are key regulatory mechanisms in cellular responses to various stimuli. The NF-jB signaling pathway is controlled by the ubiquitin-mediated proteolysis. RelA/p65, which is a main subunit of NF-jB, is ubiquitinated for degradation by SOCS-1, but the functional mechanism of its ubiquitination remains poorly understood. In this study we show that phosphorylation of RelA/p65 at Ser276 prevents its degradation by ubiquitin-mediated proteolysis. In contrast, impairment of Ser276 phosphorylation affects constitutive degradation of RelA/p65. Importantly, we identify Pim-1 as a further kinase responsible for the phosphorylation of RelA/p65 at Ser276. Depletion of Pim-1 hinders not only Ser276 phosphorylation but also transactivation of RelA/p65 target genes. We also show that Pim-1 contributes to recruitment of RelA/p65 to jB-elements to activate NF-jB signalling after TNF-a stimulation. In concert with these results, the knockdown of Pim-1 impairs IL-6 production and augments apoptosis by interfering RelA/p65 activation. These findings provide a model in which Pim-1 phosphorylation of RelA/p65 at Ser276 allows defense against ubiquitin-mediated degradation and whereby exerts activation of NF-jB signalling. NF-kB is an inducible transcription factor that controls the expression of a number of proteins involved in the regulation of cell survival and immune response. 1 It is a dimmer formed from a multisubset family consisting of RelA/p65, RelB, c-Rel, p105/p50 (NF-kB1), and p100/p52 (NF-kB2). It is activated by a bewildering array of stimuli and its activation is regulated by multiple distinct signalling cascades, including inhibitors of the NF-kB (IkB) kinase (IKK) signalosome. IKK phosphorylates IkB-a at Ser32 and Ser36 in response to a variety of stimuli, resulting in its ubiquitination and subsequent proteasomal degradation. The released NF-kB targets to the nucleus and thereby induces the expression of specific target genes. In addition to nuclear translocation of the NF-kB complex, previous studies have shown that a subunit of NF-kB, RelA/ p65, is post-translationally modified, such as phosphorylation, ubiquitination, or acetylation, and these changes influence its transcriptional activity. In particular, phosphorylation of RelA/ p65 at Ser276, Ser529, or Ser536 could be indispensable for its ability to function as an activator of gene expression. 2 However, recent findings showing a role for Ser529/536 phosphorylation on RelA/p65 activation in response to TNF-a arise conflicting evidences. [3][4][5] In contrast, accumulating evidences have revealed that Ser276 phosphorylation is critical for transactivation of RelA/p65 at least in response to TNF-a. Moreover, Ser276 is the major phosphorylation site of RelA/p65 induced by TNF-a. Phospho-RelA/p65 at Ser276 forms a stable complex with co-activator CBP/p300, a modification apparently required for assembly of a functional enhanceosome on the responsive promoters. 6,7 As nucle...

Kinase Control of Latent HIV-1 Infection: PIM-1 Kinase as a Major Contributor to HIV-1 Reactivation

“…While PIM-1 is often overexpressed in immortalized cell lines, PIM-1 is not expressed in resting primary T cells, but its expression is rapidly induced after receptor cross-linking with anti-CD3 MAbs (46). Once induced, PIM-1 has been described to phosphorylate NF-B RelA/p65 at Ser276, thereby preventing NF-B's ubiquitin-mediated proteolysis (47). PIM-1 has also been described to physically interact with NFATc1 and to phosphorylate NFATc1 in vitro on several serine residues (48).…”

NFκB–Pim-1–Eomesodermin axis is critical for maintaining CD8 T-cell memory quality

“…Thus, we hypothesized that the persistence of NFκB signaling after the peak of the response and into memory is programmed by cell-intrinsic mechanisms. Pim-1, a Ser/Thr kinase, is constitutively active once expressed and has been shown to increase p65-mediated NFκB transactivation (42,43). Most relevant to our studies, Pim-1 is expressed upon TCR stimulation.…”

Molecular Biology of Atherosclerosis

“…Phosphorylation of RelA at Ser276 is critical in facilitating RelA binding and acetylation by the coactivator CBP (CREB binding protein or closely related p300) and binding with other coactivators (such as cyclin T1/CDK9) which can direct expression of a more inflammatory subset of NF-B target genes (623,1320). Ser276 also vies for attention from the E3 ubiquitin ligase SOCS1 (suppressor of cytokine signaling 1) that targets RelA for degradation and the oncogenic kinase Pim1 (proviral integration site murine leukemia virus) that can maintain RelA active (1307). Several other posttranslational modifications are shown in FIGURE 4 (623).…”