Specificity determinants in MAPK signaling to transcription factors

One critical component in determining the specificity, fidelity, and efficiency of MAPK substrate phosphorylation is the presence of distinct docking domains in the substrate proteins. These docking domains are found in a range of substrates, including the transcription factors myocyte enhancer factor-2A and SAP-1. However, the sequences of these motifs differ, as does their targeting preferences by MAPKs, with SAP-1 being targeted by both the ERK and p38 isoforms, whereas myocyte enhancer factor-2A is targeted only by certain members of the p38 subfamily. Here, we have investigated the specificity determinants within these motifs and generated a model for how specificity is generated. We demonstrate that residues throughout the docking domains play important roles in the recognition process. However, residues located at different positions are important for discriminating between ERK and p38 MAPKs. Furthermore, the docking domains can be further subdivided into submotifs, which are differentially required for phosphorylation by ERK or p38 MAPKs. We have used loss- and gain-of-function mutagenesis to identify residues that discriminate between ERK and p38 MAPKs, residues that act to promote suboptimal interactions, and regions that are differentially required depending on the kinase involved. A model is proposed to explain how specificity is generated within these short docking domains.