Many biological processes regulated by MAPK pathways require downstream changes in gene expression. Often, this is executed by transcription factors that are direct substrates of MAPK. Capicua (Cic) is one such MAPK-regulated transcriptional repressor (1). Cic was identified as a downstream effector of receptor tyrosine kinase signaling in Drosophila, and a mammalian homolog has been identified and implicated in various cancers (2, 3). Using biophysical and genetic experiments, we aim to identify the molecular mechanisms of MAPK-mediated regulation of Cic.
MAPK pathways employ interactions between docking domains on MAPK-interacting proteins and complementary regions on MAPKs (4). We will identify a MAPK docking site on Cic. Binding of Cic to ERK2 has been mapped to a small fragment of Cic (5). However, the elements responsible for binding are unknown, and nothing is known about what part of ERK2 binds to Cic. We will mutate putative binding sites on Cic and MAPK and analyze their effects using Bio-Layer Interferometry. Initial experiments have eliminated one canonical MAPK docking site interaction as the mechanism responsible for MAPK-Cic binding. Ongoing experiments will identify whether another known docking interaction is responsible for binding or if this interaction represents a novel mechanism of MAPK-substrate recognition. Genetic experiments will introduce docking mutations into Drosophila to understand the role that binding plays in vivo. By elucidating the molecular mechanisms of Cic recognition by MAPK, we will better understand how this critical transcriptional repressor is mediated and how loss of regulation can lead to disease.