Genome-wide mapping and quantification of DNA damage induced by catechol estrogens using Click-Probe-Seq and LC-MS²

Genotoxic estrogen metabolites generate various DNA lesions; however, their target genes and carcinogenic mechanisms remain unexplored. Here, genome-wide sequencing using click probe enrichment coupled with liquid chromatography-tandem mass spectrometry (Click-Probe-Seq/LC-MS²) is developed to identify damaged genes and characterize the released and stable adducts induced by 4-hydroxy-17β-estradiol (4OHE2) in MCF-7 cell chromatin. The data reveal that guanine nucleobases in the GC-rich transcription-relevant domain are the main target sites. Moreover, the damage abundance positively correlates with DNase hypersensitive sites, suggesting that 4OHE2 preferentially attacks accessible chromatin regions beyond the estrogen receptor (ER) binding sites. Cell-based studies indicate that accumulated 4OHE2 suppresses gene transcription, causes ineffective damage repair, and decreases cell viability, differing from the uncontrolled cell growth caused by extensive ER signaling. The Click-Probe-Seq/LC-MS² approach reveals the first chromatin damage map induced by an endogenous metabolite, exposing a previously unexplored landscape in cancer research that is applicable to other genotoxic species.