PRISMA: Protein Interaction Screen on Peptide Matrix Reveals Interaction Footprints and Modifications-Dependent Interactome of Intrinsically Disordered C/EBPβ

CCAAT enhancer-binding protein beta (C/EBP beta) is a pioneer transcription factor that specifies cell differentiation. C/EBP beta is intrinsically unstructured, a molecular feature common to many proteins involved in signal processing and epigenetics. The structure of C/EBP beta differs depending on alternative translation initiation and multiple post-translational modifications (PTM). Mutation of distinct PTM sites in C/EBP beta alters protein interactions and cell differentiation, suggesting that a C/EBP beta PTM indexing code determines epigenetic outcomes. Herein, we systematically explored the interactome of C/EBP beta using an array technique based on spot-synthesized C/EBP beta-derived linear tiling peptides with and without PTM, combined with mass spectrometric proteomic analysis of protein interactions. We identified interaction footprints of similar to 1,300 proteins in nuclear extracts, many with chromatin modifying, chromatin remodeling, and RNA processing functions. The results suggest that C/EBP beta acts as a multi-tasking molecular switchboard, integrating signal-dependent modifications and structural plasticity to orchestrate interactions with numerous protein complexes directing cell fate and function.