Decoding Gene Responsiveness to Synthetic Chromatin Reader-Actuators with Multi-Modal Epigenomic Profiling

To investigate how chromatin context influences inducibility in differentiated cells, we employed engineered synthetic reader-actuators (SRAs), fusion proteins containing the polycomb chromodomain (PCD) that binds H3K27me3. In MCF7 breast cancer cells, we mapped PCD-fusion occupancy by ChIP-seq and used RNA-seq to identify temporally resolved gene activation patterns.

ChIP-seq profiling and machine learning models (MLM) demonstrated that PCD-fusion binding was predicted primarily by the absence of H3K4me3 or H3K9ac enrichment, suggesting selective accessibility at enhancers and chromatin transition zones. Among genes with SRA-enriched enhancers, the SRA-induced subset was distinguished by promoter features including bivalent histone modifications and enrichment of transcription factors NEUROD1 and MTA1.