Mitogen-activated protein kinase (MAPK)-docking sites in MAPK kinases function as tethers that are crucial for MAPK regulation in vivo.
Docking sites on targets of mitogen-activated protein kinases (MAPKs) facilitate accurate and efficient substrate phosphorylation. MAPK/ERK kinases (MEKs, or MKKs), the upstream regulators of MAPKs, also contain N-terminal MAPK-docking sites, or 'D-sites'; however, the in vivo functions of MEK D-sites are incompletely understood. Here we found that the ability of constitutively-active human MEK1 and MEK2 to stimulate ERK phosphorylation and to induce the neoplastic transformation of NIH 3T3 cells required the integrity of the D-site. In addition, D-site mutants of otherwise wild-type MEK1/2 were unable to anchor unphosphorylated ERK2 in the cytoplasm. ERK activation, cytoplasmic anchoring and release were completely retained in 'swap' mutants in which MEK2's D-site was replaced with the D-site of MEK1 or yeast Ste7. Furthermore, these abilities were significantly retained when MEK2's D-site was moved to its C-terminus, or replaced by an unrelated MAPK-binding domain taken from the Ets-1 transcription factor. We conclude that the D-sites in MEKs are crucial for the activation of their cognate MAPKs in vivo, and that their primary function is to tether their cognate MAPKs near the MEK's kinase domain. This proximity effect is sufficient to explain the contribution that the D-site interaction makes to several biologically important signaling events.
Department of Developmental and Cell Biology, 5205 McGaugh Hall, University of California Irvine, CA 92697-2300, USA.
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Grewal
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Molina
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Bardwell
D000818:Animals; D001665:Binding Sites / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000235:genetics / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology
C482128:MAP2K2 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases