Chemical profiling of DNA G-quadruplex-interacting proteins in live cells

DNA-protein interactions regulate critical biological processes. Identifying proteins that bind to specific, functional genomic loci is essential for understanding the underlying regulatory mechanisms on a molecular level. Here, we describe a novel co-binding-mediated protein profiling (CMPP) strategy to investigate the interactome of DNA G-quadruplexes (G4s) in cellular chromatin. CMPP involves cell-permeable, functionalized G4-ligand probes that bind endogenous G4s and subsequently crosslink to co-binding G4-interacting proteins in situ. We show the robustness of CMPP on proximity labelling of a G4 binding protein in vitro. Employing this approach in live cells, we identify hundreds of putative G4-interacting proteins from various functional classes. Next, we observe high G4 binding affinity and selectivity for several G4 interactors in vitro and confirm direct G4 interactions for one of the top candidates in chromatin. Our studies provide a chemical approach to map protein interactions of specific nucleic acid features in living cells. SMARCA4 ChIP-seq experiment to detect chromatin binding sites and compare to endogenous DNA G4 structures. Three technical replicates and one input.