Bacillus subtilischromosome organization oscillates between two distinct patterns

Abstract

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...

Condensin promotes the juxtaposition of DNA flanking its loading site in Bacillus subtilis

“…S1). For most of the replication-segregation cycle, the B. subtilis chromosome adopts an "ori-ter" configuration in which the left and right arms lie side by side (Wang et al 2014a). The Hi-C performed here was on asynchronously growing cells and was likely dominated by chromosomes in this configuration, accounting for the extensive interarm interactions observed.…”

“…S3A). This region spans most of the origin-proximal parS sites that are bound by ParB, forming large nucleoprotein complexes that are acted on by ParA (Murray et al 2006;Breier and Grossman 2007;Wang et al 2014a). To investigate whether these interactions require the ParABS partitioning system, we performed Hi-C on a strain lacking the parAB locus.…”

“…To investigate whether the loss of interarm interaction in the ΔparAB mutant reflects the disposition of the chromosome in single cells, we inserted tetO and lacO arrays on opposing arms of the chromosome (at ±87°) and used fluorescently tagged repressor proteins to monitor their localizations (Wang et al 2014a). We collected snapshot images of wild-type and mutant cells grown in rich and minimal media and determined interfocal distances using automated image analysis ( Fig.…”

“…ParB/parS nucleoprotein complexes are required for long-range interactions In B. subtilis, the ParABS system promotes the segregation of replicated origins (Wang et al 2014a), while ParB Figure 1. Long-range chromosome interactions require the parAB locus.…”

“…The top right panel has the color scale bar for all Hi-C contact maps presented in this study. Below the contact maps are schematics showing the ori-ter-ori configuration (Wang et al 2014a) bound to parS additionally functions to load the condensin complex in the origin region (Gruber and Errington 2009;Sullivan et al 2009;Wilhelm et al 2015). To determine which of these activities is required for the longrange interactions observed by Hi-C, we analyzed strains lacking parA, parB, or nine parS sites (eight origin-proximal parS sites at −1°, −4°, −5°, −6°, −27°, +4°, +17°, and +42°and the origin-distal site at +91°, referred to as parSΔ9) Breier and Grossman 2007).…”