Hybrid analysis reveals how DNA sequence governs genomic location and DNA contacts of bacterial chromatin H-NS filaments

Gene silencing in bacteria is mediated by chromatin proteins, of which H-NS is a paradigmatic example. H-NS forms multimeric nucleoprotein filaments with either one or two DNA duplexes, but the structures of these linear or bridged filaments remains unknown. Neither the determinants of H-NS genomic distribution nor the arrangements of H-NS DNA binding domains (DBDs) in linear or bridged filaments are currently defined. To address these questions, which are central to understanding gene silencing in bacteria, we developed a hybrid approach that combines a new method for chromatin reconstitution and mapping (Nitro-seq), ChIP-seq, nuclease-tagged DBDs to map DNA contacts, ·OH footprinting, molecular dynamics, and robust bioinformatic analyses. We find that DNA sequence principally governs genomic location of all filament types with indistinguishable sequence specificity, that DBD–DNA contacts vary in orientation and precise position with ∼10-bp spacing, and that unchanging contacts in linear and bridged filaments support a hemi-sequestration model of linear-to-bridged filament switching.