Decreased Sensitivity of Tristetraprolin-deficient Cells to p38 Inhibitors Suggests the Involvement of Tristetraprolin in the p38 Signaling Pathway

Abstract

Treatment of macrophages with pyridinyl imidazole inhibitors of p38 protein kinases can inhibit lipopolysaccharide-stimulated tumor necrosis factor ␣ secretion. However, bone marrow-derived macrophages from tristetraprolin (TTP)-deficient mice were less sensitive than normal macrophages to this effect of p38 inhibitors, despite evidence for normal p38 activation in response to lipopolysaccharide. TTP is known to cause decreased stability of tumor necrosis factor ␣ and granulocyte-macrophage colony-stimulating factor mRNAs after binding to an AU-rich element in their 3-untranslated regions. A recombinant TTP fusion protein could be phosphorylated by a recombinant p38 kinase in cellfree assays and was phosphorylated to the same extent by immunoprecipitated p38 derived from normal and TTP-deficient cells stimulated with lipopolysaccharide; in both cases, the enzyme activity was inhibited by the p38 inhibitors. TTP phosphorylation also was increased in intact macrophages after lipopolysaccharide stimulation, an effect that was blocked by the p38 inhibitors. Finally, TTP in mammalian cell extracts bound less well to an AU-rich element RNA probe than did the same amount of TTP following dephosphorylation. These results suggest that TTP may be a component of the signaling cascade, initiated by inflammatory stimuli and mediated in part by activation of p38, that ultimately leads to enhanced secretion of tumor necrosis factor ␣. Lipolysaccharide (LPS)1 -induced production of tumor necrosis factor ␣ (TNF␣) by monocyte/macrophages is regulated at both transcriptional and post-transcriptional levels. Post-transcriptional regulation of TNF␣ synthesis occurs in part by modulation of its mRNA stability. This in turn is dependent upon a so-called class II AU-rich element (ARE) found in the 3Ј-untranslated region of TNF␣ transcripts (1). This ARE has been implicated in the regulation of both TNF␣ mRNA stability and its translation (2, 3). Targeted deletion of the TNF␣ mRNA ARE in mice (⌬ARE mice) results in the overproduction of TNF␣ and the development of a systemic inflammatory syndrome (4). A role for the protein serine/threonine kinase p38 has been suggested in ARE-mediated TNF␣ mRNA processing by numerous studies (5-7), and it was found recently that macrophages from the ⌬ARE mice were relatively insensitive to the p38 inhibitor, SB203580 (4). Conflicting studies suggest that these p38 inhibitors can regulate TNF␣ synthesis at either the mRNA stability or protein translation level (8 -10). Mice lacking the p38 substrate MAPKAPK-2 have been reported to have defective TNF␣ synthesis following an LPS challenge (11). In this case, the regulation appears not to be due to a decrease in either TNF␣ mRNA levels or stability but rather to inhibition of translation, suggesting that the effects of the p38 pathway on mRNA stability and translation may be independent and uncoupled.These and other studies have indicated a role for the p38 signaling pathway in the post-transcriptional regulation of TNF␣ synthesis through a mechanism invol...

Destabilization of vascular endothelial growth factor mRNA by the zinc-finger protein TIS11b

“…In this study, the stimulation of stress-activated protein kinases induced the formation of a RNP complex that bound to a region containing the TBE and triggered stabilization of VEGF mRNA. Furthermore, it was reported that phosphorylation of TIS11 (TTP) by p38 kinase could reduce its binding affinity for ARE motifs (Carballo et al, 2001). One could then extrapolate that p38 activation might disrupt the TIS11b-TBE complex and allow recruitment of a stabilizing ARE-binding protein which remains to be identified.…”

Anti-inflammatory functions of glucocorticoid-induced genes

“…COX-2, CSF2 and IL-2 genes are dysregulated in the absence of TTP (Carballo et al, 2000;Ogilvie et al, 2005;Phillips et al, 2004), and it seems likely that expression of other inflammatory mediators will also prove to be controlled by TTP. Inflammatory mRNAs can be stabilized via the phosphorylation and inactivation of TTP by MAPK-activated protein kinase 2, a kinase that is activated by p38 MAPK (Carballo et al, 2001;Chrestensen et al, 2004;Stoecklin et al, 2004). At least in cells of the myeloid lineage, this appears to be the principal mechanism for post-transcriptional regulation of inflammatory genes by the p38 MAPK pathway.…”

Post‐transcriptional regulation of gene expression by degradation of messenger RNAs

“…TTP exists in numerous phosphorylated forms, which suggests that it may be targeted by several signaling pathways. TTP is phosphorylated by the mitogen-activated protein kinase (MAPK) p42, the p38 MAPK, and MAPK-activated protein kinase 2 (Taylor et al, 1995;Carballo et al, 2001;Mahtani et al, 2001). TTP binds to 14-3-3 proteins, the binding being largely dependent upon a specific site in the TTP C terminus, and the interaction between TTP and 14-3-3 proteins is biologically significant in determining cytoplasmic localization.…”