Mechanistic origin of cell-size control and homeostasis in bacteria

Abstract

HIGHLIGHTS• The adder requires accumulation of division proteins to a threshold for division.• The adder requires constant production of division proteins during cell elongation.• In E. coli and B. subtilis, initiation and division are independently controlled. • In E. coli and B. subtilis, cell division exclusively drives size homeostasis. GRAPHICAL ABSTRACT eTOC Blurb Si and Le Treut et al. show that cell-size homeostasis in bacteria is exclusively driven by accumulation of division proteins to a threshold and their balanced biosynthesis during cell elongation. This mechanistic insight allowed them to reprogram cell-size homeostasis in both E. coli and B. subtilis. Evolutionary implications are discussed.ABSTRACT Evolutionarily divergent bacteria share a common phenomenological strategy for cell-size homeostasis under steady-state conditions. In the presence of inherent physiological stochasticity, cells following this "adder" principle gradually return to their steady-state size by adding a constant volume between birth and division regardless of their size at birth. However, the mechanism of the adder has been unknown despite intense efforts. In this work, we show that the adder is a direct consequence of two general processes in biology: (1) threshold --accumulation of initiators and precursors required for cell division to a respective fixed number, and (2) balanced biosynthesis -maintenance of their production proportional to volume growth. This mechanism is naturally robust to static growth inhibition, but also allows us to "reprogram" cell-size homeostasis in a quantitatively predictive manner in both Gram-negative Escherichia coli and Gram-positive Bacillus subtilis. By generating dynamic oscillations in the concentration of the division protein FtsZ, we were able to oscillate cell size at division and systematically break the adder. In contrast, periodic induction of replication initiator protein DnaA caused oscillations in cell size at initiation, but did not alter division size or the adder. Finally, we were able to restore the adder phenotype in slow-growing E. coli, the only known steady-state growth condition wherein E. coli significantly deviates from the adder, by repressing active degradation of division proteins. Together these results show that cell division and replication initiation are independently controlled at the gene-expression level, and that division processes exclusively drive cell-size homeostasis in bacteria.labeled replisome protein (DnaN-YPet) to image replication cycles, and a microfluidic mother machine to follow continuous lineages during steady-state growth [3, 26] (Fig. 1A, Fig. S1; STAR Methods).A major technical challenge arises in studying replication dynamics when two replisome foci spatially overlap, which makes it difficult to analyze overlapping replication cycles. To resolve this issue, we tracked multiple replication forks from initiation to termination by extending previous imaging methods [8, 19,27,28] using the "intensity weighting" techniques [29,30] develo...

Tracking bacterial lineages in complex and dynamic environments with applications to growth control and persistence

“…Specifically, many models of growth control invoke replication patterns, and as described above, those patterns are different in entry and exit from stationary phase 39,40 . However, dividing more times without additional protein production also causes changes in the levels of division proteins, which also have been invoked in explanations of growth control 41,48 . More detailed models of how these processes change during entry into and exit from stationary phase may thus help discriminate between various hypotheses, especially if also analyzing how various mutants control growth as they enter and exit stationary phase.…”

“…For example, many growth control models invoke replication patterns, which are different in entry and exit from stationary phase 39,40 . Dividing more times without additional protein production also causes changes in the levels of division proteins, which also have been invoked in explanations of growth control 41,48 . Repeating the growth curve experiments with various genetic perturbations may thus help pinpoint the pathways involved.…”

Two different cell-cycle processes determine the timing of cell division in Escherichia coli

“…According to the first set of models cell division is controlled by DNA replication and subsequent segregation ( Witz et al, 2019; Ho and Amir, 2015; Sompayrac and Maaloe, 1973 ). According to the second set of models, cell division is controlled by a chromosome-independent inter-division process between birth and division ( Si et al, 2017, 2019; Harris and Theriot, 2016, 2018 ). C: Scheme of the concurrent-processes model.…”

“…A second class of models suggests that DNA replication has no direct influence on the timing of cell division under unperturbed growth conditions ( Harris and Theriot, 2016, 2018; Si et al, 2019; Ojkic et al, 2019; Zheng et al, 2020; Ghusinga et al, 2016 ) (Figure 1B). Instead, a different, chromosome-independent process, the accumulation of a molecule or protein, is thought to trigger cell division, once copy number reaches a threshold.…”

Dynamic Environmental Control in Microfluidic Single‐Cell Cultivations: From Concepts to Applications

“…The combination of microfluidic systems with live‐cell imaging enables dynamic cell analysis in high spatio‐temporal resolution (Figure 1D). Currently, such systems are frequently applied for investigating cell‐to‐cell heterogeneity, [ 13 ] aging and death, [ 8,13–15 ] growth, [ 8,16–18 ] cell cycle, [ 19 ] gene expression, [ 20–23 ] and various other metabolic processes. [ 24,25 ]…”