Alterations in MAPK signaling pathways can contribute to multiple diseases including Alzheimer’s disease, diabetes and cancer. Despite their high sequence similarity, the three major mammalian MAPK subfamilies (ERK1/2, JNK1/2 and p38α-δ) are activated in response to different stimuli and mediate distinct cellular responses. Short Linear Motifs (SLiMs) found in MAPK substrates and regulators, known as D-sites, are thought to play key roles in MAPK interaction specificity. These motifs bind to the D-site recruitments site (DRS), a common docking groove outside of the MAPK catalytic cleft. However, known D-site sequences conform to a common consensus sequence irrespective of its specific MAPK interaction partner. We thus lack fundamental information about how MAPKs achieve specificity. To understand how binding specificity is encoded within D-site sequences and to discover new MAPK interaction partners, we developed and executed a yeast two-hybrid screening platform to identify sequences that specifically interact with individual MAPKs. Hits selected from screening a library of 11,756 sequences from the human proteome with the MAPKs ERK2 and p38α revealed known and novel binding partners for each kinase. Alignment of the top hits uncovered strikingly distinct novel motifs selected by ERK2 and p38α. In ongoing work, we are investigating the capacity of these motifs to discriminate between MAPKs, and to investigate the structural basis for selective interactions. We are further validating novel binding partners for their capacity to act as MAPK substrates in vitro and in cells.

Specificity in the mitogen activated protein kinase signalling cascades

2019 Fall National Meeting

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