AbstractThe ChIP-exo assay precisely delineates protein-DNA crosslinking patterns by combining chromatin immunoprecipitation with 5′ to 3′ exonuclease digestion. Within a regulatory complex, the physical distance of a regulatory protein to DNA affects crosslinking efficiencies. Therefore, the spatial organization of a protein-DNA complex could potentially be inferred by analyzing how crosslinking signatures vary between the subunits of a regulatory complex. Here, we present a computational framework that aligns ChIP-exo crosslinking patterns from multiple proteins across a set of coordinately bound regulatory regions, and which detects and quantifies protein-DNA crosslinking events within the aligned profiles. By producing consistent measurements of protein-DNA crosslinking strengths across multiple proteins, our approach enables characterization of relative spatial organization within a regulatory complex. We demonstrate that our approach can recover aspects of regulatory complex spatial organization when applied to collections of ChIP-exo data that profile regulatory machinery at yeast ribosomal protein genes and yeast tRNA genes. We also demonstrate the ability to quantify changes in protein-DNA complex organization across conditions by applying our approach to data profiling Drosophila Pol II transcriptional components. Our results suggest that principled analyses of ChIP-exo crosslinking patterns enable inference of spatial organization within protein-DNA complexes.