A Novel DNA Binding Mode of H-NS Drives Chromosome Compaction and Gene Silencing in Single Bacterial Cells

Abstract

Nucleoid-associated proteins (NAPS) facilitate chromosome organization in bacteria, but the precise mechanism remains elusive. H-NS is a NAP that also plays a major role in silencing pathogen genes acquired by horizontal gene transfer. We used genetics, single-particle tracking, super-resolution fluorescence microscopy, atomic force microscopy and molecular dynamics simulations to examine H-NS/DNA interactions in single bacterial cells. We discovered a role for the unstructured linker region connecting the N-terminal oligomerization domain and the C-terminal DNA binding domain. Amino acids in the linker stabilize initial H-NS/DNA binding, facilitating polymerization of H-NS along DNA. In the absence of linker contacts, single particle tracking experiments indicated that H-NS binding was significantly reduced and in PALM images, the chromosome was de-condensed. In contrast to previous reports, H-NS was not localized to two distinct foci, rather it was scattered all around the nucleoid. Amino acids in the linker make DNA contacts that are required for both gene silencing and chromosome compaction, linking these two important functions. Supported by NIH-R21123640, VAIBX-000372 and an RCE in Mechanobiology, NUS from the Ministry of Education, Singapore.