The in vivo Interaction Landscape of Histones H3.1 and H3.3

Chromatin structure, transcription, DNA replication, and repair are regulated via locus-specific incorporation of histone variants and posttranslational modifications that guide effector chromatin-binding proteins. Here we report unbiased, quantitative interactomes for the replication-coupled (H3.1) and replication-independent (H3.3) histone H3 variants based on BioID proximity labeling, which allows interactions in intact, living cells to be detected. Along with a significant proportion of previously reported interactions detected by affinity purification followed by mass spectrometry, three quarters of the 608 histone-associated proteins that we identified are new, uncharacterized histone associations. The data reveal important biological nuances not captured by traditional biochemical means. For example, we found that the chromatin assembly factor-1 histone chaperone not only deposits the replication-coupled H3.1 histone variant during S-phase but also associates with H3.3 throughout the cell cycle in vivo. We also identified other variant-specific associations, such as with transcription factors, chromatin regulators, and with the mitotic machinery. Our proximity-based analysis is thus a rich resource that extends the H3 interactome and reveals new sets of variant-specific associations. [Display omitted] •BioID interactomes of histone variants H3.1 and H3.3.•Novel interactors and pathways were revealed.•H3, in particular H3.1, interacts with mitotic complexes.•H3.3 associates with transcription-related proteins, notably MYC interactors.•The chromatin assembly factor 1 chaperone interacts with both H3.1 and H3.3 in vivo. Proximity-dependent (BioID) interactome analysis of the histone H3 variants H3.1 and H3.3 identified 608 interaction partners and expanded the characterized H3 interaction landscape. Among the most significant findings were that the chaperone chromatin assembly factor 1, previously thought to be H3.1-specific, interacts with H3.3 throughout the cell cycle. Preferential binding was identified between H3.1 and components of the mitotic machinery and between H3.3 and transcription factors, notably MYC interactors.