Eukaryotic cells spatially organize mRNA processes such as translation and mRNA decay. Much less is clear in bacterial cells where the spatial distribution of mature mRNA remains ambiguous. Using a sensitive, quantitative fluorescence in situ hybridization based-method, we show here that in Caulobacter crescentus and Escherichia coli, chromosomally-expressed mRNAs largely display limited dispersion from their site of transcription during their lifetime. We estimate apparent diffusion coefficients at least 2 orders of magnitude lower than expected for freely diffusing mRNA, and provide evidence in C. crescentus that this mRNA localization restricts ribosomal mobility. Furthermore, C. crescentus RNase E appears associated with the DNA independently of its mRNA substrates. Collectively, our findings reveal that bacteria can spatially organize translation and potentially mRNA decay by using the chromosome layout as a template. This chromosome-centric organization has important implications for cellular physiology and for our understanding of gene expression in bacteria.

The bacterial chromosome must be compacted over 1000-fold to fit into its cellular compartment. How it is condensed, organized and ultimately segregated has been a puzzle for over half a century. Recent advances in live-cell imaging and genome-scale analyses have led to new insights into these problems. We argue that the key feature of compaction is orderly folding of DNA along adjacent segments, and that this organization provides easy and efficient access for protein-DNA transactions and plays a central role in driving segregation. Similar principles and common proteins are used in eukaryotes to condense and resolve sister chromatids at metaphase.

Summary The peptidoglycan (PG) sacculus, a meshwork of polysaccharide strands crosslinked by short peptides, protects bacterial cells against osmotic lysis. To enlarge this covalently closed macromolecule, PG hydrolases must break peptide crosslinks in the meshwork to allow insertion of new glycan strands between the existing ones. In the rod-shaped bacterium Bacillus subtilis, cell wall elongation requires two redundant endopeptidases, CwlO and LytE. However, it is not known how these potentially autolytic enzymes are regulated to prevent lethal breaches in the cell wall. Here, we show that the ATP-binding cassette transporter-like FtsEX complex is required for CwlO activity. In Escherichia coli, FtsEX is thought to harness ATP hydrolysis to activate unrelated PG hydrolases during cell division. Consistent with this regulatory scheme, B. subtilis FtsE mutants that are unable to bind or hydrolyze ATP cannot activate CwlO. Finally, we show that in cells depleted of both CwlO and LytE, the PG synthetic machinery continues moving circumferentially until cell lysis, suggesting that crosslink cleavage is not required for glycan strand polymerization. Overall, our data support a model in which the FtsEX complex is a remarkably flexible regulatory module capable of controlling a diverse set of PG hydrolases during growth and division in different organisms.

Bacterial chromosomes have been found to possess one of two distinct patterns of spatial organization. In the first, called "ori-ter" and exemplified by Caulobacter crescentus, the chromosome arms lie side-by-side, with the replication origin and terminus at opposite cell poles. In the second, observed in slow-growing Escherichia coli ("left-ori-right"), the two chromosome arms reside in separate cell halves, on either side of a centrally located origin. These two patterns, rotated 90°relative to each other, appear to result from different segregation mechanisms. Here, we show that the Bacillus subtilis chromosome alternates between them. For most of the cell cycle, newly replicated origins are maintained at opposite poles with chromosome arms adjacent to each other, in an ori-ter configuration. Shortly after replication initiation, the duplicated origins move as a unit to midcell and the two unreplicated arms resolve into opposite cell halves, generating a left-ori-right pattern. The origins are then actively segregated toward opposite poles, resetting the cycle. Our data suggest that the condensin complex and the parABS partitioning system are the principal driving forces underlying this oscillatory cycle. We propose that the distinct organization patterns observed for bacterial chromosomes reflect a common organization-segregation mechanism, and that simple modifications to it underlie the unique patterns observed in different species.DNA replication | ParA | chromosome segregation | SMC condensin C entral to reproduction is the faithful segregation of replicated chromosomes to daughter cells. In eukaryotes, DNA replication, chromosome condensation, and sister chromatid segregation are separated into distinct steps in the cell cycle that are safeguarded by checkpoint pathways. In bacteria, these processes occur concurrently, posing unique challenges to genome integrity and inheritance (1, 2). In the absence of temporal control, bacteria take advantage of spatial organization to promote faithful and efficient chromosome segregation. The organization of the chromosome dictates where the chromosome is replicated, and the factors that organize and compact the newly replicated DNA play a central role in its segregation (1, 2).Studies in different bacteria have revealed strikingly distinct patterns of chromosome organization that appear to arise from different segregation mechanisms. In Caulobacter crescentus and Vibrio cholerae chromosome I, the origin and terminus are located at opposite cell poles, with the two replication arms between them, in a pattern referred to as "ori-ter" (3-5). After replication initiation, one of the sister origins is held in place and the other is actively translocated to the opposite cell pole, regenerating the oriter organization in both daughter cells (5-11). By contrast, in slowgrowing Escherichia coli, the origin is located in the middle of the nucleoid and the two replication arms reside in opposite cell halves, in a "left-ori-right" pattern (12, 13). Replication initiates at mid...