A conserved docking site in MEKs mediates high-affinity binding to MAP kinases and cooperates with a scaffold protein to enhance signal transmission.
The recognition of mitogen-activated protein kinases (MAPKs) by their upstream activators, MAPK/ERK kinases (MEKs), is crucial for the effective and accurate transmission of many signals. We demonstrated previously that the yeast MAPKs Kss1 and Fus3 bind with high affinity to the N terminus of the MEK Ste7, and proposed that a conserved motif in Ste7, the MAPK-docking site, mediates this interaction. Here we show that the corresponding sequences in human MEK1 and MEK2 are necessary and sufficient for the direct binding of the MAPKs ERK1 and ERK2. Mutations in MEK1, MEK2, or Ste7 that altered conserved residues in the docking site diminished binding of the cognate MAPKs. Furthermore, short peptides corresponding to the docking sites in these MEKs inhibited MEK1-mediated phosphorylation of ERK2 in vitro. In yeast cells, docking-defective alleles of Ste7 were modestly compromised in their ability to transmit the mating pheromone signal. This deficiency was dramatically enhanced when the ability of the Ste5 scaffold protein to associate with components of the MAPK cascade was also compromised. Thus, both the MEK-MAPK docking interaction and binding to the Ste5 scaffold make mutually reinforcing contributions to the efficiency of signaling by this MAPK cascade in vivo.
Bardwell
A J
AJ
Department of Developmental and Cell Biology, University of California, Irvine, California 92697, USA. bardwell@uci.edu
Flatauer
L J
LJ
Matsukuma
K
K
Thorner
J
J
Bardwell
L
L
D048868:Adaptor Proteins, Signal Transducing; D000483:Alleles; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins; D017124:Conserved Sequence; D004305:Dose-Response Relationship, Drug; D005656:Fungal Proteins / Q000378:metabolism; D005800:Genes, Fungal; D005982:Glutathione Transferase / Q000378:metabolism; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000737:chemistry / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D016366:Open Reading Frames; D010455:Peptides / Q000737:chemistry; D010675:Pheromones / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D011494:Protein Kinases / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014158:Transcription, Genetic
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D005656:Fungal Proteins; D010455:Peptides; D010675:Pheromones; D011993:Recombinant Fusion Proteins; C080321:STE5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D005982:Glutathione Transferase; D011494:Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C119844:STE7 protein, S cerevisiae