Dynamic bistable switches enhance robustness and accuracy of cell cycle transitions

Bistability is a common mechanism to ensure robust and irreversible cell cycle transitions. Whenever biological parameters or external conditions change such that a threshold is crossed, the system abruptly switches between different cell cycle states. Experimental studies have uncovered mechanisms that can make the shape of the bistable response curve change dynamically in time. Here, we show how such a dynamically changing bistable switch can provide a cell with better control over the timing of cell cycle transitions. Moreover, cell cycle oscillations built on bistable switches are more robust when the bistability is modulated in time. Our results are not specific to cell cycle models and may apply to other bistable systems in which the bistable response curve is time-dependent.

Response to comment on ‘Initiation of chromosome replication controls both division and replication cycles in E. coli through a double-adder mechanism’

Last year we published an article (Witz et al., 2019) in which we used time-lapse microscopy in combination with microfluidics to measure growth, division and replication in single E. coli cells on the one hand, and developed a new statistical analysis method to calculate the ability of different cell cycle models to capture the correlation structure observed in the data on the other hand. This led us to propose a new model of cell cycle control in E. coli which we called the double-adder model.Recently Le Treut et al. published a comment (Le Treut et al., 2020) on our article which made a number of highly critical claims, including allegations that our own data support a different model than the one we proposed, and that our model cannot reproduce the ‘adder phenotype’ observed in the data. We here show that all these allegations are false and based on basic analysis errors. Although our focus is on explaining the errors in the analysis of Le Treut et al, we have attempted to make the presentation of interest to a broader scientific audience by discussing the issues in the context of what our current understanding is of the bacterial cell cycle, and to what extent recent data either support or reject various proposed models.

Initiation of chromosome replication controls both division and replication cycles in E. coli through a double-adder mechanism

Living cells proliferate by completing and coordinating two cycles, a division cycle controlling cell size and a DNA replication cycle controlling the number of chromosomal copies. It remains unclear how bacteria such as Escherichia coli tightly coordinate those two cycles across a wide range of growth conditions. Here, we used time-lapse microscopy in combination with microfluidics to measure growth, division and replication in single E. coli cells in both slow and fast growth conditions. To compare different phenomenological cell cycle models, we introduce a statistical framework assessing their ability to capture the correlation structure observed in the data. In combination with stochastic simulations, our data indicate that the cell cycle is driven from one initiation event to the next rather than from birth to division and is controlled by two adder mechanisms: the added volume since the last initiation event determines the timing of both the next division and replication initiation events.

One and two-phase cell cycle models

In this review paper we present deterministic and stochastic one and two-phase models of the cell cycle. The deterministic models are given by partial differential equations of the first order with time delay and space variable retardation. The stochastic models are given by stochastic iterations or by piecewise deterministic Markov processes. We study asymptotic stability and sweeping of stochastic semigroups which describe the evolution of densities of these processes. We also present some results concerning chaotic behaviour of models and relations between different types of models.

Control of the Restriction Point by Rb and p21

The Restriction Point was originally defined as the moment that cells commit to the cell cycle and was later suggested to coincide with hyperphosphorylation of the retinoblastoma protein (Rb). Current cell cycle models posit that cells exit mitosis into a pre-Restriction Point state, where they have low cyclin-dependent kinase (CDK) activity and hypophosphorylated Rb; passage through the Restriction Point then occurs in late G1. Recent single-cell studies have challenged the current paradigm, raising questions about the location of the Restriction Point and the notion that cells exit mitosis into a pre-Restriction Point state. Here, we use a variety of single-cell techniques to show that both noncancer and cancer cells bifurcate into two subpopulations after anaphase, marked by increasing vs. low CDK2 activity and hyper- vs. hypophosphorylation of Rb. Notably, subpopulations with hyper- and hypophosphorylated Rb are present within minutes after anaphase, delineating one subpopulation that never “uncrosses” the Restriction Point and continues cycling and another subpopulation that exits mitosis into an uncommitted pre-Restriction Point state. We further show that the CDK inhibitor p21 begins rising in G2 in mother cells whose daughters exit mitosis into the pre-Restriction Point, CDK2low state. Furthermore, degradation of p21 coincides with escape from the CDK2low state and passage through the Restriction Point. Together, these data support a model in which only a subset of cells returns to a pre-Restriction Point state after mitosis and where the Restriction Point is sensitive to not only mitogens, but also inherited DNA replication stress via p21.

Novel chromosome organization pattern in actinomycetales–overlapping replication cycles combined with diploidy

Bacteria regulate chromosome replication and segregation tightly with cell division to ensure faithful segregation of DNA to daughter generations. The underlying mechanisms have been addressed in several model species. It became apparent that bacteria have evolved quite different strategies to regulate DNA segregation and chromosomal organization. We have investigated here how the actinobacteriumCorynebacterium glutamicumorganizes chromosome segregation and DNA replication. Unexpectedly, we find thatC. glutamicumcells are at least diploid under all conditions tested and that these organisms have overlapping C-periods during replication with both origins initiating replication simultaneously. Based on experimentally obtained data we propose growth rate dependent cell cycle models forC. glutamicum.