Spatiotemporal and global profiling of DNA–protein interactions enables discovery of low-affinity transcription factors

Precise dissection of DNA–protein interactions is essential for elucidating the recognition basis, dynamics and gene regulation mechanism. However, global profiling of weak and dynamic DNA–protein interactions remains a long-standing challenge. Here, we establish the light-induced lysine (K) enabled crosslinking (LIKE-XL) strategy for spatiotemporal and global profiling of DNA–protein interactions. Harnessing unique abilities to capture weak and transient DNA–protein interactions, we demonstrate that LIKE-XL enables the discovery of low-affinity transcription-factor/DNA interactions via sequence-specific DNA baits, determining the binding sites for transcription factors that have been previously unknown. More importantly, we successfully decipher the dynamics of the transcription factor subproteome in response to drug treatment in a time-resolved manner, and find downstream target transcription factors from drug perturbations, providing insight into their dynamic transcriptional networks. The LIKE-XL strategy offers a complementary method to expand the DNA–protein profiling toolbox and map accurate DNA–protein interactomes that were previously inaccessible via non-covalent strategies, for better understanding of protein function in health and disease.