Epigenetic competition reveals density-dependent regulation and target site plasticity of phosphorothioate epigenetics in bacteria.

Phosphorothioate (PT) DNA modifications—in which a nonbonding phosphate oxygen is replaced with sulfur—represent a widespread, horizontally transferred epigenetic system in prokaryotes and have a highly unusual property of occupying only a small fraction of available consensus sequences in a genome. Using Salmonella enterica as a model, we asked a question of fundamental importance: How do the PT-modifying DndA-E proteins select their GPSAAC/GPSTTC targets? Here, we applied innovative analytical, sequencing, and computational tools to discover a novel behavior for DNA-binding proteins: The Dnd proteins are “parked” at the G6mATC Dam methyltransferase consensus sequence instead of the expected GAAC/GTTC motif, with removal of the 6mA permitting extensive PT modification of GATC sites. This shift in modification sites further revealed a surprising constancy in the density of PT modifications across the genome. Computational analysis showed that GAAC, GTTC, and GATC share common features of DNA shape, which suggests that PT epigenetics are regulated in a density-dependent manner partly by DNA shape-driven target selection in the genome.