MKK3- and MKK6-Regulated Gene Expression Is Mediated by the p38 Mitogen-Activated Protein Kinase Signal Transduction Pathway

Abstract

The p38 mitogen-activated protein (MAP) kinase signal transduction pathway is activated by proinflammatory cytokines and environmental stress. The detection of p38 MAP kinase in the nucleus of activated cells suggests that p38 MAP kinase can mediate signaling to the nucleus. To test this hypothesis, we constructed expression vectors for activated MKK3 and MKK6, two MAP kinase kinases that phosphorylate and activate p38 MAP kinase. Expression of activated MKK3 and MKK6 in cultured cells caused a selective increase in p38 MAP kinase activity. Cotransfection experiments demonstrated that p38 MAP kinase activation causes increased reporter gene expression mediated by the transcription factors ATF2 and Elk-1. These data demonstrate that the nucleus is one target of the p38 MAP kinase signal transduction pathway.Several mitogen-activated protein (MAP) kinase signal transduction pathways have been detected in mammalian cells (15). Three groups of MAP kinases have been molecularly cloned: ERK (7, 8), JNK (16,22,31,34,55), and p38 (27,35,47). These MAP kinases are activated by dual phosphorylation on Thr and Tyr within the motif Thr-Xaa-Tyr in subdomain VIII (15). The sequence of this dual phosphorylation motif differs for each MAP kinase group as follows: p38, Thr-GlyTyr; JNK, Thr-Pro-Tyr; and ERK, Thr-Glu-Tyr. Each MAP kinase group has a distinct substrate specificity and is regulated by a separate signal transduction pathway (15). Mammalian cells therefore contain multiple MAP kinase signal transduction pathways that mediate the effects of extracellular stimuli on a wide array of biological processes.Detailed studies of the JNK and ERK groups of MAP kinase have led to significant insight into the physiological function of these signaling pathways (6, 13-15, 40, 45). In contrast, the role of the p38 MAP kinase signal transduction pathway is poorly understood (20,27,35,44,47). p38 MAP kinase is weakly activated by protein kinase C and receptor tyrosine kinases but is strongly activated by the treatment of cells with inflammatory cytokines (e.g., tumor necrosis factor and interleukin-1) and environmental stress (e.g., osmotic shock and UV radiation) (20,27,35,44,47). The contribution of the p38 MAP kinase pathway to the cellular response to these stimuli has not been established. However, recent studies have implicated p38 MAP kinase in the phosphorylation of the small heat shock protein Hsp27 (20, 47), in increased cytokine expression (35), and in programmed cell death (61). Furthermore, in vitro protein kinase assays demonstrate that p38 MAP kinase phosphorylates MAPKAP kinase-2 (20, 47) and the transcription factor ATF2 (17, 44).The mechanism of p38 MAP kinase activation is mediated by dual phosphorylation on Thr and Tyr within the motif ThrGly-Tyr located in subdomain VIII (44). The p38 MAP kinase activator MKK3 has been molecularly cloned (17). MKK3 is a protein kinase that phosphorylates and activates p38 MAP kinase but does not phosphorylate the related JNK or ERK MAP kinases (17). MKK3 is therefore a specific act...

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