In Escherichia coli crosstalk between DNA supercoiling, nucleoid-associated proteins and major RNA polymerase σ initiation factors regulates growth phase-dependent gene transcription. We show that the highly conserved spatial ordering of relevant genes along the chromosomal replichores largely corresponds both to their temporal expression patterns during growth and to an inferred gradient of DNA superhelical density from the origin to the terminus. Genes implicated in similar functions are related mainly in trans across the chromosomal replichores, whereas DNA-binding transcriptional regulators interact predominantly with targets in cis along the replichores. We also demonstrate that macrodomains (the individual structural partitions of the chromosome) are regulated differently. We infer that spatial and temporal variation of DNA superhelicity during the growth cycle coordinates oxygen and nutrient availability with global chromosome structure, thus providing a mechanistic insight into how the organization of a complete bacterial chromosome encodes a spatiotemporal program integrating DNA replication and global gene expression. In Escherichia coli cells the physiological transitions induced by the changing growth environment are accompanied by changes in DNA superhelical density (1-3), nucleoid structure (4-6), and the promoter selectivity of the RNA polymerase (RNAP) holoenzyme (3, 7). During the growth cycle both the relative and absolute concentrations of the abundant nucleoid-associated proteins (NAPs ; Table S1) change substantially and correspondingly generate bacterial chromatin of variable composition (8, 9). The NAPs stabilize distinct supercoil structures (10-12) selectively favoring particular RNAP holoenzymes (13-15). These variable nucleoprotein complexes modulate DNA topology during the growth cycle (Fig. 1A), optimizing the channeling of supercoil energy into appropriate metabolic pathways (16,17).The expression of the genes determining superhelical density, polymerase selectivity, and nucleoid structure is coordinated by cross-regulation. Thus, factor for inversion stimulation (FIS), a NAP abundant during the early exponential phase (18), regulates expression not only of the superhelicity determinants DNA gyrase subunits A and B (gyrA and gyrB) and topoisomerase I (topA) (19-21) but also other NAP-encoding genes including hns, α subunit of histone-like protein from E. coli strain U93 (hupA), and DNA binding protein from starved cells (dps) (22-24) and components of the transcription machinery such as σ 38 subunit of RNA polymerase rpoS (25). Similarly mutations affecting the selectivity of RNAP influence NAP production (26-28). Again, mutations in the genes controlling DNA superhelicity affect the production of both NAPs and the basal transcription machinery (27,29). This pattern of integrated control constitutes a heterarchical network coordinating chromosome structure with cellular metabolism (27,28,30). A further pointer to this integrated network is the observed selection of mutations in fis...

The chromosomal DNA polymer constituting the cellular genetic material is primarily a device for coding information. Whilst the gene sequences comprise the digital (discontinuous) linear code, physiological alterations of the DNA superhelical density generate in addition analog (continuous) three-dimensional information essential for regulation of both chromosome compaction and gene expression. Insight into the relationship between the DNA analog information and the digital linear code is of fundamental importance for understanding genetic regulation. Our previous study in the model organism Escherichia coli suggested that the chromosomal gene order and a spatiotemporal gradient of DNA superhelicity associated with DNA replication determine the growth phase-dependent gene transcription. In this study we reveal a general gradient of DNA thermodynamic stability correlated with the polarity of chromosomal replication and manifest in the spatiotemporal pattern of gene transcription during the bacterial growth cycle. Furthermore, by integrating the physical and dynamic features of the transcribed sequences with their functional content we identify spatiotemporal domains of gene expression encompassing different functions. We thus provide both an insight into the organisational principle of the bacterial growth program and a novel holistic methodology for exploring chromosomal dynamics.

Recent studies strongly suggest that the gene expression sustaining both normal and pathogenic bacterial growth is governed by the structural dynamics of the chromosome. However, the mechanistic device coordinating the chromosomal configuration with selective expression of the adaptive traits remains largely unknown. We used a holistic approach exploring the inherent relationships between the physicochemical properties of the DNA and the expression of adaptive traits, including virulence factors, in the pathogen Dickeya dadantii (formerly Erwinia chrysanthemi). In the transcriptomes obtained under adverse conditions encountered during bacterial infection, we explored the patterns of chromosomal DNA sequence organization, supercoil dynamics, and gene expression densities, together with the long-range regulatory impacts of the abundant DNA architectural proteins implicated in pathogenicity control. By integrating these data, we identified transient chromosomal domains of coherent gene expression featuring distinct couplings between DNA thermodynamic stability, supercoil dynamics, and virulence traits.

Over the recent decade, the central importance of DNA supercoiling in chromosome organization and global gene regulation of bacteria became more and more visible. With a regulon comprising more than 2000 genes in Escherichia coli, DNA supercoiling is among the most influential regulators of gene expression found in bacteria so far. However, the mechanism creating thousands of diverse temporal gene expression patterns coordinated by DNA supercoiling remains unclear. In this study we show that a specific chromosomal arrangement of genes modulates the local levels of DNA supercoiling at gene promoters via transcription-coupled DNA supercoiling (TCDS) in the model organism E. coli. Our findings provide a consistent explanation for the strong positive coupling of temporal gene expression patterns of neighboring genes. Using comparative genomics we are furthermore able to provide evidence that TCDS is a driving force for the evolution of chromosomal gene arrangement patterns in other Enterobacteriaceae. With the currently available data of promoter supercoiling sensitivity we prove that the same principle is applicable also for the evolutionary distant gram-positive pathogenic bacterium Streptococcus pneumoniae. Moreover, our findings are fully consistent with recent investigations concerning the regulatory impact of TCDS on gene pairs in eukaryots underpinning the broad applicability of our analysis.

A considerable share of bacterial species maintains multipartite genomes. Precise coordination of genome replication and segregation with cell growth and division is vital for proliferation of these bacteria. The α-proteobacterium Sinorhizobium meliloti possesses a tripartite genome composed of one chromosome and the megaplasmids pSymA and pSymB. Here, we investigated the spatiotemporal pattern of segregation of these S. meliloti replicons at single cell level. Duplication of chromosomal and megaplasmid origins of replication occurred spatially and temporally separated, and only once per cell cycle. Tracking of FROS (fluorescent repressor operator system)-labelled origins revealed a strict temporal order of segregation events commencing with the chromosome followed by pSymA and then by pSymB. The repA2B2C2 region derived from pSymA was sufficient to confer the spatiotemporal behaviour of this megaplasmid to a small plasmid. Altering activity of the ubiquitous prokaryotic replication initiator DnaA, either positively or negatively, resulted in an increase in replication initiation events or G1 arrest of the chromosome only. This suggests that interference with DnaA activity does not affect replication initiation control of the megaplasmids.