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