Mapping the human proteome with physical access to DNA

In a human cell, DNA is packed in histones, RNA, and chromatin-associated proteins, forming a cohesive gel. At any given moment, only a specific subset of the proteome has physical access to the DNA and organizes its structure, transcription, replication, repair and other molecular functions essential to the way the genome is read and maintained. We have developed a "zero-distance" photo-crosslinking approach to quantify proteins in direct contact with DNA in living cells. Collecting DNA interactomes from human breast cancer cells, we present an atlas of over one thousand proteins with physical access to DNA, and hundreds of peptide-nucleotide crosslinks pinpointing protein-DNA interfaces with single amino-acid resolution. Differential comparisons of DNA interactomes from cells undergoing treatment with estrogen or genotoxic chemotherapy recapitulated the recruitment of key transcription factors and DNA damage proteins. This opens a direct way to explore genomic regulation in a hypothesis-free manner, applicable to many organisms and systems.