Comprehensive chromatin proteomics resolves functional phases of pluripotency

The establishment of cellular identity is driven by transcriptional and epigenetic regulation exerted by the components of the chromatin proteome - the chromatome. However, chromatome composition and its dynamics in functional phases of pluripotency have not been comprehensively analyzed thus limiting our understanding of these processes. To address this problem, we developed an accurate mass spectrometry (MS)-based proteomic method called Chromatin Aggregation Capture (ChAC) followed by Data-Independent Acquisition (DIA) to analyze chromatome reorganizations during the transition from ground to formative and primed pluripotency states. This allowed us to generate a comprehensive atlas of proteomes, chromatomes, and chromatin affinities for the three pluripotency phases, revealing the specific binding and rearrangement of regulatory complexes. The technical advances, the comprehensive chromatome atlas, and the extensive analysis reported here provide a foundation for an in-depth understanding of mechanisms that govern the phased progression of pluripotency and changes of cellular identities in development and disease.