Metabolites such as short-chain fatty acids can interact with chromatin and affect transcription by modifying histone proteins. The variety of histone modifications makes it technically hard to delineate their function in a site- and modification-type-specific way. Qin et al. developed a single-site-resolved multi-omics (SiTomics) strategy based on genetic code expansion technology to profile the interacting proteomic and genomic landscape for a given histone modification in cells. The authors first used an unbiased proteomics approach to identify histone-modification sites in response to short-chain fatty acids, revealing known modification sites (such as K56) and new sites (K36, K37 and K64) on histone H3. To characterize these modifications under physiological conditions, the authors used genetic code expansion to incorporate lysine analogs that carry a given modification into a specific site of histone H3 to generate pre-modified histones in living cells, which could then be integrated into chromatin for genomic location reading via next-generation sequencing. The photoaffinity moiety in the lysine analogs enables the formation of covalent crosslinking with proximal proteins after light illumination, and the crosslinked proteins can be identified through mass spectrometry. Using their SiTomics strategy, the authors showed that unique modifications even at the same histone site can have diverse interactomes and gene-regulation functions. For example, H3K56 crotonylation showed enrichment in promoter regions and exhibited a cooperative effect on facilitating nucleosomal DNA accessibility with the active histone mark H3K4me3. H3K56 β-hydroxybutrylation can be enriched at super-enhancers, along with H3K27acetylation and transcriptional coactivators, resulting in looser 3D chromatin organization. These results indicate that SiTomics is a powerful platform for elucidating site-specific functions of histone modifications in a physiologically relevant environment.
Original reference: Cell 186, 1066–1085 (2023).
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