Tiny but Mighty: How do Bacteria Maintain Their Size?
When we think about cellular diversity, one feature that easily comes to mind is cellular size. An illustration of this is through the bacterial kingdom: Escherichia coli (E. coli) averages at 1-2 µm in length, while Candidatus Thiomargarita magnifica measures in a centimetre (Volland et al., 2022). Classically, changes to cell size is viewed as a function of nutrient availability, as explained by the Growth Law (Cesar and Huang, 2017). Yet, at steady state, cell size remains constant as observed in Bacillus subtilis, despite time extending beyond its doubling duration (Levin and Angert, 2015). Similarly, earlier experiments probing into the mystery of cell size pointed to a level of regulation. Scientists Hartmann and Prescott reduced the size of an amoeba by experimentally micro-dissecting part of its cytoplasm and drew remarkable observations (Rhind, 2021). In the experimentally smaller amoeba, cell division was halted until its size increased to normal (Rhind, 2021). Hartmann, in a valiant attempt, microdissected an amoeba’s cytoplasm daily for four months thereby, halting cell division for approximately 65 cycles; yet, the amoeba grew to its normal size and underwent normal cell division (Rhind, 2021). Overall, these earlier observations suggest that a target size must be met prior to cell division, implying a layer of regulation must exist.
Essentially, the central question becomes: how does a single cell or a unicellular organism, such as bacteria, decide they are big enough? In this article, we will explore the current understanding underlying bacterial cell size control.
The adder model
The prevailing model explaining bacterial size homeostasis is known as the “adder” model. This model proposes that during each cell cycle event, a fixed volume is added to the cell regardless of its size at birth (Sauls et al., 2016). As symmetric cell division and population growth proceeds, sizes of newborn cells will gradually converge to the average size of the population (Figure 1) (Amir, 2014; Taheri-Araghi et al., 2015). Direct experimental evidence of adder behaviour was obtained through interdisciplinary approaches of single-cell time lapse microscopy and mathematical modelling (Campos et al., 2014; Taheri-Araghi et al., 2015). For example, tracking of E.coli at single-cell resolution over seven hours revealed that on average, E.coli cells grew the same length before undergoing cell division (Campos et al., 2014). Furthermore, no correlation was found between the length of growth with the initial cell size at birth (Campos et al., 2014), thus providing support for the adder model.
Figure 1. Schematic of E.coli size convergence towards the population size average through the adder model. Cells born larger (top) or smaller (bottom) than the population average (dashed line) correct their size over successive generations by adding a constant size increase (Δ) prior to cell division, independent of their initial size at birth. This leads to an exponential convergence towards the mean cell size, stabilising the population’s size distribution over multiple division cycles. Figure adapted from Sauls et al. (2016).
Sizing things up, at a molecular level
While these previous works have helped set the stage in experimentally validating the adder model, how size homeostasis is controlled at a molecular level remains unknown. To understand this, researchers have hypothesised that size regulation begins at the onset of DNA replication (replication-initiation view) or cell division (division-centric view) (Sauls et al., 2016). A general principle shared by both hypotheses is that cells monitor their size by accumulating levels of key regulatory proteins to certain thresholds which triggers either DNA replication or cell division.
The rationale for the replication-initiation view is drawn from observations that DNA replication is initiated when E.coli are of the same size and mass (Donachie, 1968; Si et al., 2017). However, E.coli cells which are 30% smaller relative to normal, are unable to undergo division despite being able to initiate DNA replication (Grossman and Ron, 1989). Furthermore, several studies have shown that shorter E.coli cells maintain similar growth rates as normal cells, however modulate its cell cycle duration to achieve size homeostasis (Campos et al., 2014; Lambert et al., 2018). Thus, whether size regulation begins at the initiation of DNA replication or cell division remains debated.
To investigate both possibilities, Si et al. (2019) experimentally modulated expression levels of DnaA, a protein vital in replication initiation in E.coli (Si et al., 2019). By oscillating dnaA expression, E.coli was found to initiate DNA replication at different sizes while the overall growth rate remained unchanged (Si et al., 2019). This highlights the role of DnaA accumulation as a proxy for the cell to gauge its size during growth, and trigger replication once a critical threshold is met. Interestingly, all cells divided at normal size, and increased size at the onset of division was only observed when timing of replication termination overlapped with cell division (Si et al., 2019). This suggests size homeostasis may instead be independently controlled by cell division events. In support of this, Si et al. (2019) modulated the expression levels of FtsZ, a tubulin-like protein that forms a ring at the cell’s midpoint to enable the assembly of the cell division machinery (Cameron and Margolin, 2024). Unlike during DnaA level modulation, oscillating ftsZ expression levels affect cellular size at division, and is triggered by the total accumulation of FtsZ (Si et al., 2019). As such, cell division events regulate a cell’s size by determining how big of a cell is considered “too big”, while also regulating and establishing the size of its daughter cells at birth.
A major limitation of these studies is that these experiments were conducted under steady-state conditions, which do not accurately reflect the fluctuating environmental conditions which bacteria face in nature. Computational models in recent years have helped to bridge this gap, providing potential insights. For example, Serbanescu et al. (2020) developed a theoretical framework demonstrating that under nutrient-poor conditions, average cellular size is reduced as cells redirect ribosomal resources to synthesise division proteins such as FtsZ, over promoting growth (Serbanescu et al., 2020). As nutrient availability improves, cell size increases as growth becomes re-prioritised, leading to larger cell sizes (Serbanescu et al., 2020), and recent quantitative studies have further validated this (Kratz and Banerjee, 2023).
Conclusion
Although Prescott and Hartmann’s pioneering experiments date back several decades, the intricate mechanisms underlying cell size control have only been unveiled in the past decade. Largely, this progress is attributed to the advent of interdisciplinary approaches and experimental tools that have enabled interrogation of bacterial behaviour at a single-cell level. While much remains to be learned, it remains exciting to witness the elegant complexity of these homeostatic mechanisms.
Article prepared by: Maxine Tan, MBIOS R&D Associate 24/25
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References
Volland, Jean-Marie. "Small cells with big secrets." Nature Reviews Microbiology 21.7 (2023): 414-414.
Cesar, Spencer, and Kerwyn Casey Huang. "Thinking big: the tunability of bacterial cell size." FEMS microbiology reviews 41.5 (2017): 672-678.
Levin, Petra Anne, and Esther R. Angert. "Small but mighty: cell size and bacteria." Cold Spring Harbor perspectives in biology 7.7 (2015): a019216.
Rhind, Nicholas. "Cell-size control." Current Biology 31.21 (2021): R1414-R1420.
Sauls, John T., Dongyang Li, and Suckjoon Jun. "Adder and a coarse-grained approach to cell size homeostasis in bacteria." Current opinion in cell biology 38 (2016): 38-44.
Amir, Ariel. "Cell size regulation in bacteria." Physical review letters 112.20 (2014): 208102.
Taheri-Araghi, Sattar, et al. "Cell-size control and homeostasis in bacteria." Current biology 25.3 (2015): 385-391.
Campos, Manuel, et al. "A constant size extension drives bacterial cell size homeostasis." Cell 159.6 (2014): 1433-1446.
Donachie, William D. "Relationship between cell size and time of initiation of DNA replication." Nature 219.5158 (1968): 1077-1079.
Si, Fangwei, et al. "Invariance of initiation mass and predictability of cell size in Escherichia coli." Current Biology 27.9 (2017): 1278-1287.
Grossman, N. I. L. I., and ELIORA Z. Ron. "Apparent minimal size required for cell division in Escherichia coli." Journal of bacteriology 171.1 (1989): 80-82.
Lambert, Ambroise, et al. "Constriction rate modulation can drive cell size control and homeostasis in C. crescentus." Iscience 4 (2018): 180-189.
Si, Fangwei, et al. "Mechanistic origin of cell-size control and homeostasis in bacteria." Current
Biology 29.11 (2019): 1760-1770.
Cameron, Todd A., and William Margolin. "Insights into the assembly and regulation of the bacterial divisome." Nature Reviews Microbiology 22.1 (2024): 33-45.
Serbanescu, Diana, Nikola Ojkic, and Shiladitya Banerjee. "Nutrient-dependent trade-offs between ribosomes and division protein synthesis control bacterial cell size and growth." Cell reports 32.12 (2020).
Kratz, Josiah C., and Shiladitya Banerjee. "Dynamic proteome trade-offs regulate bacterial cell size and growth in fluctuating nutrient environments." Communications Biology 6.1 (2023): 486.
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