Chromosome segregation in B. subtilis is highly heterogeneous
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RESEARCH NOTE
Chromosome segregation in B. subtilis is highly heterogeneous Nina El Najjar1,2 and Peter L. Graumann1,2*
Abstract Objective: The bacterial cell cycle comprises initiation of replication and ensuing elongation, concomitant chromosome segregation (in some organisms with a delay termed cohesion), and finally cell division. By quantifying the number of origin and terminus regions in exponentially growing Bacillus subtilis cells, and after induction of DNA damage, we aimed at determining cell cycle parameters at different growth rates at a single cell level. Results: B. subtilis cells are mostly mero-oligoploid during fast growth and diploid during slow growth. However, we found that the number of replication origins and of termini is highly heterogeneous within the cell population at two different growth rates, and that even at slow growth, a majority of cells attempts to maintain more than a single chromosome at all times of the cell cycle. Heterogeneity of chromosome copy numbers may reflect different subpopulations having diverging growth rates even during exponential growth conditions. Cells continued to initiate replication and segregate chromosomes after induction of DNA damage, as judged by an increase in origin numbers per cell, showing that replication and segregation are relatively robust against cell cycle perturbation. Keywords: Bacterial cell cycle, Chromosome segregation, Bacillus subtilis, Single cell analyses Introduction The segregation of chromosomes is a central part of the cell cycle in all kinds of cells, yet poorly understood in bacteria [1, 2]. In a previous study, cell cycle parameters of Bacillus subtilis have been characterized by determination of population averages of DNA content and nucleoid morphology at four different growth rates. The pattern of increased numbers of replication forks at higher growth rates was similar compared to Escherichia coli cells: at low growth rates, B. subtilis was mostly mono or diploid (later in the cell cycle), meaning cells had one chromosome copy or two, while cells became mero-oligoploid at higher growth rates [3], meaning they contain several origin regions per chromosome copy because of overlapping round of replication. More recent publications have
shown that the situation is different from what was calculated, namely that few cells are monoploid, even in poor growth medium [4], and that during exponential phase, most cells are mero-oligoploid in rich medium, and reduced chromosome copy numbers during entry into stationary phase [5]. In our recent study, we have shown that the pattern of chromosome segregation follows that of a linear, directed manner, and could be explained by entropy driven segregation based on diffusion [6]. In this work, we extend these analyses by scoring the number of origin or terminus regions, visualized by FROS, at two different growth rates, namely minimal and rich medium. We also characterized chromosome copy numbers by quantitative DAPI staining of the DNA, and investigated c
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