ERK MAP kinase is targeted to RSK2 by the phosphoprotein PEA-15

Abstract

The ERK pathway responds to extracellular stimuli and oncogenes by modulating cellular processes, including transcription, adhesion, survival, and proliferation. ERK has diverse substrates that carry out these functions. The processes that are modulated are determined in part by the substrates that ERK phosphorylates. We demonstrate that PEA-15 (phosphoprotein enriched in astrocytes, 15 kDa) targets ERK to RSK2 and thereby enhances RSK2 activation. PEA-15 independently bound ERK and RSK2 and increased ERK association with RSK2 in a concentration-dependent manner. PEA-15 increased RSK2 activity and CREB-mediated transcription, and this process was regulated by phosphorylation of PEA-15. Finally, phorbol ester stimulation of PEA-15-null lymphocytes resulted in impaired RSK2 activation that was rescued by exogenous PEA-15 expression. Therefore, PEA-15 functions as a scaffold to enhance ERK activation of RSK2, and this activity is regulated by phosphorylation. Thus, PEA-15 can integrate signal transduction to provide a specific physiological outcome from activation of the multipotent ERK MAP kinase pathway.signal transduction ͉ scaffold ͉ transcription ͉ lymphocytes T he ERK MAPK signaling cascade has been implicated in diverse physiological processes, including differentiation, proliferation, migration, and adhesion (1). This ability to impact several processes is partly because ERK recognizes an array of substrates both cytoplasmic and nuclear to elicit specific responses. However, how such a multipotent pathway achieves specificity of response is an ongoing problem in understanding ERK function. ERK interaction with scaffolding proteins, regulation of the duration of the ERK signal, and restriction of ERK to specialized subcellular compartments have all been suggested to contribute to this specificity (2-4). However, thus far, the mammalian scaffolding proteins for ERK such as KSR are known to act only upstream of ERK to enhance ERK activation without directing ERK to a particular substrate (5, 6). PEA-15 (phosphoprotein enriched in astrocytes, 15 kDa) is a small death effector domain protein that binds ERK and RSK2 and sequesters them in the cytoplasm (7,8). RSK2 is a substrate of ERK and a kinase that can activate transcription, regulate apoptosis, and control cancer cell proliferation (9-11). Therefore, we investigated whether PEA-15 influences ERK activation of RSK2. We show that increasing expression of PEA-15 enhances ERK binding to RSK2, ERK phosphorylation of RSK2, and RSK2 activity. Moreover, genetic deletion of PEA-15 substantially abrogates ERK activation of RSK2 in astrocytes and lymphocytes. RSK2 activation can be rescued by ectopic expression of PEA-15 in these cells. Thus, PEA-15 functions as a scaffold to enhance ERK binding and phosphorylation of RSK2. It does not affect the phosphorylation of other ERK substrates, such as stathmin and Mnk1. This work provides a model to understand how scaffold proteins can integrate signal transduction and provide specificity in MAP kinase signaling.

Mitochondrial hexokinase II (HKII) and phosphoprotein enriched in astrocytes (PEA15) form a molecular switch governing cellular fate depending on the metabolic state

“…Similar to HKII (17), up-regulation of PEA15 is an important feature of multiple malignant tumors (39). The mitochondrial localization of PEA15 reported above was unexpected because it has previously been described as a cytosolic protein, interacting with Fas-associated protein with death Domain (FADD) and FADD-like IL-1β-converting enzyme (FLICE) through its death effector domain (DED), thereby blocking the formation of the death-inducing signaling complex (DISC) (41). However, PEA15 mediates sensitivity to Bcl2-induced survival in cells undergoing mitochondria-dependent type-II apoptosis (42) and also interacts with the mitochondrial serine protease Omi/HtrA2 (43).…”

“…PEA15 is involved in the regulation of apoptosis, glucose metabolism, cellular proliferation, and oncogenesis (26,(37)(38)(39)(40)(41), and it has been suggested to contribute to insulin resistance in type 2 diabetes (26). Similar to HKII (17), up-regulation of PEA15 is an important feature of multiple malignant tumors (39).…”

Phospholipase C∈ Activates Nuclear Factor-κB Signaling by Causing Cytoplasmic Localization of Ribosomal S6 Kinase and Facilitating Its Phosphorylation of Inhibitor κB in Colon Epithelial Cells

“…The association of RSK with PEA15 seemed independent of its Ser-380 phosphorylation. Moreover, LPA stimulation elevated the phosphorylation of PEA15 at Ser-104, a putative PKC target site implicated in enhancing the cytoplasmic localization of RSK, which was also enhanced by PLC⑀ overexpression (33) (Fig. 7E).…”

“…We focused on PEA15, a scaffold protein for RSK in the cytoplasm, which had been reported to regulate subcellular localization of RSK, depending on the phosphorylation state (32,33). RSK could be co-immunoprecipitated with PEA15 in Caco2 cells stimulated with LPA, which was markedly enhanced by PLC⑀ overexpression (Fig.…”

“…The subcellular localization of RSK is reported to be regulated by a scaffold protein, PEA15 (phosphoprotein enriched in astrocytes 15), which directly associates with RSK and holds it in the cytoplasm, based on the observation that overexpression of PEA15 prevented nuclear translocation of RSK (32). Moreover, overexpression of a phosphorylation-mimicking mutant, S104D, of PEA15 failed to elevate the CREB promoter activity, suggesting that the phosphorylation of PEA15 might regulate the RSK localization (33).…”