MK2 and MK3 represent protein kinases downstream of p38 mitogen-activated protein kinase (MAPK).Deletion of the MK2 gene in mice resulted in an impaired inflammatory response although MK3, which displays extensive structural similarities and identical functional properties in vitro, is still present. Here, we analyze tumor necrosis factor (TNF) production and expression of p38 MAPK and tristetraprolin (TTP) in MK3-deficient mice and demonstrate that there are no significant differences with wild-type animals. We show that in vivo MK2 and MK3 are expressed and activated in parallel. However, the level of activity of MK2 is always significantly higher than that of MK3. Accordingly, we hypothesized that MK3 could have significant effects only in an MK2-free background and generated MK2/MK3 double-knockout mice. Unexpectedly, these mice are viable and show no obvious defects due to loss of compensation between MK2 and MK3. However, there is a further reduction of TNF production and expression of p38 and TTP in double-knockout mice compared to MK2-deficient mice. This finding, together with the observation that ectopically expressed MK3 can rescue MK2 deficiency similarly to MK2, indicates that both kinases share the same physiological function in vivo but are expressed to different levels.Downstream of mitogen-activated protein kinases (MAPKs) different groups of MAPK-activated protein kinases (MAP KAPKs) have been defined (reviewed in reference 28). These enzymes transduce signals to target proteins that are not direct substrates of the MAPKs and, therefore, serve to relay phosphorylation-dependent signaling within MAPK cascades to diverse cellular functions. One of these groups is formed by the three MAPKAPKs, MK2, MK3 (also known as 3pK), and MK5 (also designated PRAK) (reviewed in reference 12). While MK5 is mainly activated by the atypical MAPK ERK3 (29, 30), the remaining two kinases, MK2 and MK3, are directly downstream of the MAPK p38␣/␤ (7,10,24,27,31). Phosphorylation of MK2 and MK3 by p38␣/␤ at two or three major regulatory sites leads to activation and coupled nuclear export of both enzymes, which are localized in the nucleus of resting cells (4,8,26,36,41).A wide variety of substrates has been described for MK2 including proteins interacting with the cytoskeleton, such as small heat shock protein Hsp25 (33); mRNA-binding proteins, such as tristetraprolin (TTP) (6, 32); transcription factors, such as heat shock factor 1 (38); and regulators of the cell cycle and apoptosis, such as Cdc25B/C (23). The phosphorylation site recognition motifs of MK2 and MK3 are similar (20) or even identical (7). Despite the similar recognition motif, not all MK2 substrates have been described as MK3 substrates so far, probably because in most cells MK2 activity dominates and makes analysis of the minor MK3 activity dependent on antibodies which discriminate between both enzymes (7).MK2-deficient mice are more resistant than wild type to endotoxic shock due to impaired production of cytokines such as tumor necrosis factor (T...

TNF expression of macrophages is under stringent translational control that depends on the p38 MAPK/MK2 pathway and the AU–rich element (ARE) in the TNF mRNA. Here, we elucidate the molecular mechanism of phosphorylation-regulated translation of TNF. We demonstrate that translation of the TNF-precursor at the ER requires expression of the ARE–binding and -stabilizing factor human antigen R (HuR) together with either activity of the p38 MAPK/MK2 pathway or the absence of the ARE-binding and -destabilizing factor tristetraprolin (TTP). We show that phosphorylation of TTP by MK2 decreases its affinity to the ARE, inhibits its ability to replace HuR, and permits HuR-mediated initiation of translation of TNF mRNA. Since translation of TTP's own mRNA is also regulated by this mechanism, an intrinsic feedback control of the inflammatory response is ensured. The phosphorylation-regulated TTP/HuR exchange at target mRNAs provides a reversible switch between unstable/non-translatable and stable/efficiently translated mRNAs.

Receptor-interacting protein kinase-1 (RIPK1), a master regulator of cell fate decisions, was identified as a direct substrate of MAPKAP kinase-2 (MK2) by phosphoproteomic screens using LPS-treated macrophages and stress-stimulated embryonic fibroblasts. p38/MK2 interact with RIPK1 in a cytoplasmic complex and MK2 phosphorylates mouse RIPK1 at Ser321/336 in response to pro-inflammatory stimuli, such as TNF and LPS, and infection with the pathogen Yersinia enterocolitica. MK2 phosphorylation inhibits RIPK1 autophosphorylation, curtails RIPK1 integration into cytoplasmic cytotoxic complexes, and suppresses RIPK1-dependent apoptosis and necroptosis. In Yersinia-infected macrophages, RIPK1 phosphorylation by MK2 protects against infection-induced apoptosis, a process targeted by Yersinia outer protein P (YopP). YopP suppresses p38/MK2 activation to increase Yersinia-driven apoptosis. Hence, MK2 phosphorylation of RIPK1 is a crucial checkpoint for cell fate in inflammation and infection that determines the outcome of bacteria-host cell interaction.

Most dendritic cell (DC) responses to Toll-like receptor (TLR) ligands depend on the activation of mitogen-activated protein kinases (MAPKs), but the contributions of the many MAPK-activated kinases (MKs) that act 'downstream' of the MAPKs Erk and p38 are not known. Here we sought to determine which MKs are required for acute TLR-driven, MAPK-dependent DC endocytic responses. Two specific and structurally different inhibitors of the MK Rsk suppressed TLR-induced endocytosis, thus defining in DCs a specific requirement for MKs in TLR responses. In addition, we identify in DCs a previously unknown configuration of the MAPK system whereby Rsk is activated not only by Erk but also by p38 through the intermediates MK2 and MK3. Thus, in DCs, p38 contributes to the activation of all known MK families.

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