Condensation of nucleoid during environmental adaptation is regulated by dynamic HU interactions

Nucleoid remodeling facilitated by DNA supercoiling results in changes in nucleoid configuration and involves nucleoid-associated proteins (NAPs) and structural maintenance of chromosomes (SMC) proteins among others. Changes in nucleoid configurations regulated by NAPs are synchronized with cellular adaptation and influence the simultaneous expression of several genes. HU, a ubiquitous bacterial histone-like protein, is among the most conserved and abundant NAPs in eubacteria. In Escherichia coli, HU forms dimers by HUa self-association (HUaa) or by HUa-HUß interactions (HUaß). HUa is mostly expressed during lag and early exponential growth phase and HUß is expressed only during the later exponential and stationary phase, pointing to distinct HUaa/DNA and HUaß/DNA packaging of the nucleoid in regulating expression patterns during growth and stasis. Mutations or the deletion of HU transform the E. coli nucleoid to a different form and alter overall transcription program; thus, HU interactions with DNA directly affect global gene regulation. Recently, analysis of high-resolution contact maps of the E. coli nucleoid revealed an important role of HU to promote long-range DNA-DNA contacts within the nucleoid. Yet, the molecular connections between HU-DNA interactions and changes in nucleoid architecture that regulate gene expression globally remain unknown. Here, we explored the higher-order E. coli nucleoid organization by soft x-ray tomography (SXT) and revealed an effect of HU surface charges in overall nucleoid organization and rearrangements. We also studied global transcription by next-generation RNA sequencing (RNA-Seq) and found a link between nucleoid rearrangement and changes in global transcription. To determine the functional relationships of observed nucleoid rearrangement and HU-DNA interactions, we also characterized the overall organization of HU nucleoprotein complexes in solution by small angle x-ray scattering (SAXS). We found that HUaa-mediated DNA networks are different at different ionic strengths and pHs. By means of macromolecular crystallography, we additionally elucidate HUaa dependent molecular switches that modulate DNA networking. This integrative structural study explains how HUaa regulates dynamic transformations of the nucleoid by DNA bridging to control nucleoid rearrangement and global gene regulation. Overall design: Three biological replicates were analyzed for each of 12 conditions