An Efficient Approach for Selective Enrichment of Histone Modification Readers Using Self-Assembled Multivalent Photoaffinity Peptide Probes.

Histone post-translational modifications (HPTMs) provide signaling platforms to recruit proteins or protein complexes (e.g., transcription factors, the so-called "readers" of the histone code), changing DNA accessibility in the regulation of gene expression. Thus, it is an essential task to identify HPTM readers for understanding of epigenetic regulation. Herein we designed and prepared a novel HPTM probe based on self-assembled multivalent photo-cross-linking technique for selective enrichment and identification of HPTM readers. By use of trimethylation of histone H3 lysine 4, we showcased that the functionalized HPTM probe was able to capture its reader with high enrichment efficiency and remarkable specificity even in a complex environment. Notably, this approach was readily applicable for exploring crosstalk among multiple HPTMs. Combining the probes with a mass spectrometry-based proteomic approach, our approach reached a fairly high coverage of known H3K4me3 readers. We further demonstrated that the HPTM probes can enrich a new type of HPTM readers and uncovered several novel putative binders of crotonylation of histone H3 lysine 9, expanding the repertoire of readers for this epigenetic mark. More broadly, our work provides a general strategy for rapid and robust interrogating HPTM readers and will be of great importance to elucidate epigenetic mechanism in regulating gene activity.