Biology subjects

ten Wolde, P. R.

Publications and source records attributed to ten Wolde, P. R..

9 recordsLinked to original sources

bioRxiv2026

Growth, metabolism, and the benefits of clock-enabled anticipation

Circadian clocks are found in myriad species across the tree of life. They are commonly assumed to provide a selective advantage by enabling organisms to anticipate daily environmental changes. It has proven remarkably difficult, however, to move beyond this general idea to explain precisely what advantage anticipation confers and to quantify how large this benefit is. Here, we address these challenges by developing a mathematical model of diurnally varying growth and metabolism in the cyanobacterium Synechococcus elongatus, which fixes carbon through photosynthesis during the day and stores a portion of it as glycogen for use during the night. Specifically, we extend the framework introduced by Hwa and coworkers (Scott et al., Science 330, 1099 (2010)), which quantifies the relationship between growth and proteome composition, to a circadian context. We find that the fastest growth is achieved when glycogen production during the day is minimized to the amount required to sustain essential nighttime processes; growth then occurs exclusively during the day, in agreement with experimental observations. However, the slow relaxation of the proteome, arising from the intrinsically slow growth rate of S. elongatus, severely constrains the implementation of this optimal strategy: In the absence of anticipation, the cell must grow at night in order to produce proteins essential for nighttime metabolism. In contrast, with anticipation enabled by a circadian clock, the bacterium can instead adjust its proteome late in the day, allowing it to avoid less efficient nighttime growth altogether, thereby significantly enhancing its growth rate.

microbiology↗

bioRxiv2026

Establishment of titration-based control of DNA replication in Escherichia coli

Escherichia coli couples the initiation of DNA replication with cell size by modulating the activity of the replication initiator protein DnaA. The activity of DnaA is regulated by both its interconversion between an active and inactive form and its titration on binding sites on the chromosome. Whereas its interconversion has been thoroughly studied, the extent to which DnaA titration can control replication initiation is poorly understood. Here, we describe the control of E. coli DNA replication via titration by modulating the expression of an always-active DnaA variant in four growth conditions. While we obtained stable cell cycles during slow growth, faster growth associated with overlapping replication forks led to replicative instability and DNA damage. Overall, our results provide insights into the limits of titration-based systems in the control of genome replication and their potential role in the evolutionary trajectory of E. coli. Finally, this study provides design principles for a simplified, titration-only regulatory mechanism for DNA replication in synthetic cells.

molecular biology↗

bioRxiv2024

Quantifying the nuclear localisation of fluorescently tagged proteins

Cells are highly dynamic, continually responding to intra- and extracellular signals. Monitoring and measuring the response to these signals at the single-cell level is challenging. Signal transduction is fast, but reporters for downstream gene expression are typically slow, requiring fluorescent proteins to be expressed and to mature. An alternative is to fluorescently tag and then monitor the intracellular locations of transcription factors and other effectors. These proteins move in or out of the nucleus in minutes, after upstream signalling modifies their state of phosphorylation. Although such approaches are being used increasingly, there is no consensus on how to quantify the nuclear and cytoplasmic localisation of these proteins. Using budding yeast, we developed a convolutional neural network that quantifies nuclear localisation from fluorescence and, optionally, bright-field images. Focusing on the cellular response to changing glucose, we generated ground-truth data using strains with both a transcription factor and a nuclear protein tagged with fluorescent markers. We then showed that the neural network based approach outperformed seven published methods, particularly when predicting single-cell time series, which are key to determining how cells respond and adapt. Collectively, our results are conclusive -- using machine learning to automatically determine the appropriate image processing consistently outperforms ad hoc methods. Adopting such methods promises to both improve the accuracy and, with transfer learning, the consistency of single-cell analyses.

bioinformatics↗

bioRxiv2024

The Escherichia coli replication initiator DnaA is titrated on the chromosome

DNA replication initiation is orchestrated in many prokaryotes by the replication initiator DnaA. Two models for regulation of DnaA activity in Escherichia coli have been proposed: the switch between an active and inactive form of DnaA, and the titration of DnaA on the E. coli chromosome. Although proposed decades ago, experimental evidence of a titration-based control mechanism is still lacking. Here, we first identified a conserved high-density region of binding motifs near the origin of replication, an advantageous trait for titration of DnaA. We then investigated the mobility of DnaA by single-particle tracking microscopy in wild-type and deletion mutants E. coli strains, while monitoring cellular size and DNA content. Our results indicate that the chromosome of E. coli controls the free amount of DnaA in a growth rate-dependent fashion. Finally, we provide insights on the relevance of DnaA titration in stabilising DNA replication by preventing re-initiation events during slow growth.

molecular biology↗

bioRxiv2023

Trade-offs between cost and information in cellular prediction

Living cells can leverage correlations in environmental fluctuations to predict the future environment and mount a response ahead of time. To this end, cells need to encode the past signal into the output of the intracellular network from which the future input is predicted. Yet, storing information is costly while not all features of the past signal are equally informative on the future input signal. Here, we show, for two classes of input signals, that cellular networks can reach the fundamental bound on the predictive information as set by the information extracted from the past signal: pushpull networks can reach this information bound for Markovian signals, while networks that take a temporal derivative can reach the bound for predicting the future derivative of non-Markovian signals. However, the bits of past information that are most informative about the future signal are also prohibitively costly. As a result, the optimal system that maximizes the predictive information for a given resource cost is, in general, not at the information bound. Applying our theory to the chemotaxis network of Escherichia coli reveals that its adaptive kernel is optimal for predicting future concentration changes over a broad range of background concentrations, and that the system has been tailored to predicting these changes in shallow gradients.

biophysics↗

bioRxiv2022

Robust replication initiation from coupled homeostatic mechanisms

The bacterium Escherichia coli initiates replication once per cell cycle at a precise volume per origin and adds an on average constant volume between successive initiation events, independent of the initiation size. Yet, a molecular model that can explain these observations has been lacking. Experiments indicate that E. coli controls replication initiation via titration and activation of the initiator protein DnaA. Here, we study by mathematical modelling how these two mechanisms interact to generate robust replication-initiation cycles. We first show that a mechanism solely based on titration generates stable replication cycles at low growth rates, but inevitably causes premature reinitiation events at higher growth rates. In this regime, the DnaA activation switch becomes essential for stable replication initiation. Conversely, while the activation switch alone yields robust rhythms at high growth rates, titration can strongly enhance the stability of the switch at low growth rates. Our analysis thus predicts that both mechanisms together drive robust replication cycles at all growth rates. In addition, it reveals how an origin-density sensor yields adder correlations.

biophysics↗

bioRxiv2021

Cross-linkers at growing microtubule ends generate forces that drive actin transport

The actin and microtubule cytoskeletons form active networks in the cell that can contract and remodel, resulting in vital cellular processes as cell division and motility. Motor proteins play an important role in generating the forces required for these processes, but more recently the concept of passive cross-linkers being able to generate forces has emerged. So far, these passive cross-linkers have been studied in the context of separate actin and microtubule systems. Here, we show that crosslinkers also allow actin and microtubules to exert forces on each other. More specifically, we study single actin filaments that are cross-linked to growing microtubule ends, using in vitro reconstitution, computer simulations, and a minimal theoretical model. We show that microtubules can transport actin filaments over large (micrometer-range) distances, and find that this transport results from two antagonistic forces arising from the binding of cross-linkers to the overlap between the actin and microtubule filaments. The cross-linkers attempt to maximize the overlap between the actin and the tip of the growing microtubules, creating an affinity-driven forward condensation force, and simultaneously create a competing friction force along the microtubule lattice. We predict and verify experimentally how the average transport time depends on the actin filament length and the microtubule growth velocity, confirming the competition between a forward condensation force and a backward friction force. In addition, we theoretically predict and experimentally verify that the condensation force is of the order of 0.1 pN. Thus, our results reveal a new mechanism for local actin remodeling by growing microtubules. Significance StatementComplex cellular processes such as cell migration require coordinated remodeling of both the actin and the microtubule cytoskeleton. The two networks for instance exert forces on each other via active motor proteins. Here we show that, surprisingly, coupling via passive cross-linkers can also result in force generation. We specifically study the transport of actin filaments by growing microtubule ends. We show by cell-free reconstitution experiments, computer simulations, and theoretical modeling that this transport is driven by the affinity of the cross-linker for the chemically distinct microtubule tip region. Our work predicts that growing microtubules could potentially rapidly relocate newly nucleated actin filaments to the leading edge of the cell and thus boost migration.

biophysics↗

bioRxiv2018

Frustrated binding of biopolymer crosslinkers

Transiently crosslinked actin filament networks allow cells to combine elastic rigidity with the ability to deform viscoelastically. Theoretical models of semiflexible polymer networks predict that the crosslinker unbinding rate governs the timescale beyond which viscoelastic flow occurs. However a direct comparison between network and crosslinker dynamics is lacking. Here we measure the networks stress relaxation timescale using rheology and the lifetime of bound crosslinkers using fluorescence recovery after photobleaching. lntruigingly, we observe that the crosslinker unbinding rate measured by FRAP is more than an order of magnitude slower than the rate measured by rheology. We rationalize this difference with a three-state model where crosslinkers are bound to either 0, 1 or 2 filaments, which allows us to extract crosslinker transition rates that are otherwise difficult to access. We find that the unbinding rate of singly bound crosslinkers is nearly two orders of magnitude slower than for double bound ones. We attribute the increased unbinding rate of doubly bound crosslinkers to the high stiffness of biopolymers, which frustrates crosslinker binding.

biophysics↗

bioRxiv2018

Theory of Circadian Metabolism

Many organisms repartition their proteome in a circadian fashion in response to the daily nutrient changes in their environment. A striking example is provided by cyanobacteria, which perform photosynthesis during the day to fix carbon. These organisms not only face the challenge of rewiring their proteome every 12 hours, but also the necessity of storing the fixed carbon in the form of glycogen to fuel processes during the night. In this manuscript, we extend the framework developed by Hwa and coworkers (Scott et al., Science 330, 1099 (2010)) for quantifying the relatinship between growth and proteome composition to circadian metabolism. We then apply this framework to investigate the circadian metabolism of the cyanobacterium Cyanothece, which not only fixes carbon during the day, but also nitrogen during the night, storing it in the polymer cyanophycin. Our analysis reveals that the need to store carbon and nitrogen tends to generate an extreme growth strategy, in which the cells predominantly grow during the day, as observed experimentally. This strategy maximizes the growth rate over 24 hours, and can be quantitatively understood by the bacterial growth laws. Our analysis also shows that the slow relaxation of the proteome, arising from the slow growth rate, puts a severe constraint on implementing this optimal strategy. Yet, the capacity to estimate the time of the day, enabled by the circadian clock, makes it possible to anticipate the daily changes in the environment and mount a response ahead of time. This significantly enhances the growth rate by counteracting the detrimental effects of the slow proteome relaxation.

systems biology↗