Cell Cycle Oscillator Research Articles

The malaria parasite has an intrinsic clock.

Malarial rhythmic fevers are the consequence of the synchronous bursting of red blood cells (RBCs) on completion of the malaria parasite asexual cell cycle. Here, we hypothesized that an intrinsic clock in the parasite Plasmodium chabaudi underlies the 24-hour-based rhythms of RBC bursting in mice. We show that parasite rhythms are flexible and lengthen to match the rhythms of hosts with long circadian periods. We also show that malaria rhythms persist even when host food intake is evenly spread across 24 hours, suggesting that host feeding cues are not required for synchrony. Moreover, we find that the parasite population remains synchronous and rhythmic even in an arrhythmic clock mutant host. Thus, we propose that parasite rhythms are generated by the parasite, possibly to anticipate its circadian environment.

Nonequilibrium Thermodynamics in Cell Biology: Extending Equilibrium Formalism to Cover Living Systems.

We discuss new developments in the nonequilibrium dynamics and thermodynamics of living systems, giving a few examples to demonstrate the importance of nonequilibrium thermodynamics for understanding biological dynamics and functions. We study single-molecule enzyme dynamics, in which the nonequilibrium thermodynamic and dynamic driving forces of chemical potential and flux are crucial for the emergence of non-Michaelis-Menten kinetics. We explore single-gene expression dynamics, in which nonequilibrium dissipation can suppress fluctuations. We investigate the cell cycle and identify the nutrition supply as the energy input that sustains the stability, speed, and coherence of cell cycle oscillation, from which the different vital phases of the cell cycle emerge. We examine neural decision-making processes and find the trade-offs among speed, accuracy, and thermodynamic costs that are important for neural function. Lastly, we consider the thermodynamic cost for specificity in cellular signaling and adaptation.

Clb3-centered regulations are recurrent across distinct parameter regions in minimal autonomous cell cycle oscillator designs

Some biological networks exhibit oscillations in their components to convert stimuli to time-dependent responses. The eukaryotic cell cycle is such a case, being governed by waves of cyclin-dependent kinase (cyclin/Cdk) activities that rise and fall with specific timing and guarantee its timely occurrence. Disruption of cyclin/Cdk oscillations could result in dysfunction through reduced cell division. Therefore, it is of interest to capture properties of network designs that exhibit robust oscillations. Here we show that a minimal yeast cell cycle network is able to oscillate autonomously, and that cyclin/Cdk-mediated positive feedback loops (PFLs) and Clb3-centered regulations sustain cyclin/Cdk oscillations, in known and hypothetical network designs. We propose that Clb3-mediated coordination of cyclin/Cdk waves reconciles checkpoint and oscillatory cell cycle models. Considering the evolutionary conservation of the cyclin/Cdk network across eukaryotes, we hypothesize that functional (“healthy”) phenotypes require the capacity to oscillate autonomously whereas dysfunctional (potentially “diseased”) phenotypes may lack this capacity.

Cell cycle oscillators underlying orderly proteolysis of E2F8.

E2F8 is a transcriptional repressor that antagonizes E2F1 at the crossroads of the cell cycle, apoptosis, and cancer. Previously, we discovered that E2F8 is a direct target of the APC/C ubiquitin ligase. Nevertheless, it remains unknown how E2F8 is dynamically controlled throughout the entirety of the cell cycle. Here, using newly developed human cell-free systems that recapitulate distinct inter-mitotic and G1 phases and a continuous transition from prometaphase to G1, we reveal an interlocking dephosphorylation switch coordinating E2F8 degradation with mitotic exit and the activation of APC/CCdh1. Further, we uncover differential proteolysis rates for E2F8 at different points within G1 phase, accounting for its accumulation in late G1 while APC/CCdh1 is still active. Finally, we demonstrate that the F-box protein Cyclin F regulates E2F8 in G2-phase. Altogether, our data define E2F8 regulation throughout the cell cycle, illuminating an extensive coordination between phosphorylation, ubiquitination and transcription in mammalian cell cycle.

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 indicate that the shape of the bistable response curve changes 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.

C-di-GMP Arms an Anti-σ to Control Progression of Multicellular Differentiation in Streptomyces.

SummaryStreptomyces are our primary source of antibiotics, produced concomitantly with the transition from vegetative growth to sporulation in a complex developmental life cycle. We previously showed that the signaling molecule c-di-GMP binds BldD, a master repressor, to control initiation of development. Here we demonstrate that c-di-GMP also intervenes later in development to control differentiation of the reproductive hyphae into spores by arming a novel anti-σ (RsiG) to bind and sequester a sporulation-specific σ factor (σWhiG). We present the structure of the RsiG-(c-di-GMP)2-σWhiG complex, revealing an unusual, partially intercalated c-di-GMP dimer bound at the RsiG-σWhiG interface. RsiG binds c-di-GMP in the absence of σWhiG, employing a novel E(X)3S(X)2R(X)3Q(X)3D motif repeated on each helix of a coiled coil. Further studies demonstrate that c-di-GMP is essential for RsiG to inhibit σWhiG. These findings reveal a newly described control mechanism for σ-anti-σ complex formation and establish c-di-GMP as the central integrator of Streptomyces development.

Inference of the High-Level Interaction Topology between the Metabolic and Cell-Cycle Oscillators from Single-Cell Dynamics.

Recent evidence suggests that the eukaryotic metabolism is an autonomous oscillator. Together with oscillating elements of the cyclin/CDK machinery, this oscillator might form a coupled oscillator system, from which cell-cycle control emerges. The topology of interactions between the metabolic oscillator and the elements of the cyclin/CDK machinery, however, remains unknown. Using single-cell metabolic and cell-cycle dynamics in yeast, and solving an inverse problem with a system of Kuramotooscillators, we inferred how the metabolic oscillator interacts with the cyclin/CDK machinery. The identified and experimentally validated interaction topology shows that the early and late cell cycle are independently driven by metabolism. While in this topology, the S phase is coordinated by START. We obtained no support for a strong interaction between early and late cell cycle. The identified high-level interaction topology will guide future efforts to discover the molecular links between metabolism and the cell cycle.

Process-induced cell cycle oscillations in CHO cultures: Online monitoring and model-based investigation.

The influence of process strategies on the dynamics of cell population heterogeneities in mammalian cell culture is still not well understood. We recently found that the progression of cells through the cell cycle causes metabolic regulations with variable productivities in antibody-producing Chimese hamster ovary (CHO) cells. On the other hand, it is so far unknown how bulk cultivation conditions, for example, variable nutrient concentrations depending on process strategies, can influence cell cycle-derived population dynamics. In this study, process-induced cell cycle synchronization was assessed in repeated-batch and fed-batch cultures. An automated flow cytometry set-up was developed to measure the cell cycle distribution online, using antibody-producing CHO DP-12 cells transduced with the cell cycle-specific fluorescent ubiquitination-based cell cycle indicator (FUCCI) system. On the basis of the population-resolved model, feeding-induced partial self-synchronization was predicted and the results were evaluated experimentally. In the repeated-batch culture, stable cell cycle oscillations were confirmed with an oscillating G1 phase distribution between 41% and 72%. Furthermore, oscillations of the cell cycle distribution were simulated and determined in a (bolus) fed-batch process with up to cells/ml. The cell cycle synchronization arose with pulse feeding only and ceased with continuous feeding. Both simulated and observed oscillations occurred at higher frequencies than those observable based on regular (e.g., daily) sample analysis, thus demonstrating the need for high-frequency online cell cycle analysis. In summary, we showed how experimental methods combined with simulations enable the improved assessment of the effects of process strategies on the dynamics of cell cycle-dependent population heterogeneities. This provides a novel approach to understand cell cycle regulations, control cell population dynamics, avoid inadvertently induced oscillations of cell cycle distributions and thus to improve process stability and efficiency.

A Mathematical Model for the Effect of Low-Dose Radiation on the G2/M Transition.

We develop a mathematical model to study the immediate effect of low-dose radiation on the G2 checkpoint and the G2/M transition of the cell cycle via a radiation pathway (the ATM-Chk2 pathway) of an individual mammalian cell. The model consists of a system of nonlinear differential equations describing the dynamics of a network of regulatory proteins that play key roles in the G2/M transition, cell cycle oscillations, and the radiation pathway. We simulate the application of a single pulse of low-dose radiation at different intensities ([Formula: see text]0-0.4Gy) and times during the latter part of the G2-phase. We use bifurcation analysis to characterize the effect of radiation on the G2/M transition via the ATM-Chk2 pathway. We show that radiation between 0.1 and 0.3Gy can delay the G2/M transition, and radiation higher than 0.3Gy can fully activate the G2 checkpoint. Also, our results show that radiation can be low enough to neither delay the G2/M transition nor activate the G2 checkpoint ([Formula: see text]0.1Gy). Our model supports the idea that the cell response to radiation during G2-phase explains hyper-radiosensitivity and increased radioresistance (HRS/IRR) observed at low dose.

Building Dynamic Cellular Machineries in Droplet-Based Artificial Cells with Single-Droplet Tracking and Analysis.

Although the application of droplet microfluidics has grown exponentially in chemistry and biology over the past decades, robust universal platforms for the routine generation and comprehensive analysis of droplet-based artificial cells are still rare. Here we report using microfluidic droplets to reproduce a variety of types of cellular machinery in in vitro artificial cells. In combination with a unique image-based analysis method, the system enables full automation in tracking single droplets with high accuracy, high throughput, and high sensitivity. These powerful performances allow broad applicability evident in three representative droplet-based analytical prototypes that we develop for (i) droplet digital detection, (ii) in vitro transcription and translation reactions, and (iii) spatiotemporal dynamics of cell-cycle oscillations. The capacities of this platform to generate, incubate, track, and analyze individual microdroplets via real-time, long-term imaging unleash its great potential in accelerating cell-free synthetic biology. Moreover, the wide scope covering from digital to analog to morphological detections makes this droplet analysis technique adaptable for many other divergent types of droplet-based chemical and biological assays.

Spontaneous Synchronisation and the Cell Cycle

We suggest that it may be possible for the cell cycles of neighboring biological cells to become spontaneously synchronized, e.g., in the presence of defects or disease. We are not proposing that spontaneous synchronization of cycles between neighboring cells is necessarily common or permanent. Rather, we suggest that temporary islands or “domains” of cells with synchronized cell cycles can form and exhibit collective behavior for some time, before disappearing. We suggest that a cell with faulty DNA whose cycle is synchronized with those of surrounding healthy cells may be dragged past the G1/S and G2/M checkpoints even if, in isolation, it would be stalled to allow repair or marking for apoptosis. Synchronisation may occur if cell cycle oscillations are coupled with receipt and transmission of messages at cell membranes. Storms of inter-cell messages could cause collaborative cell cycling and synchronised cell growth, which may in turn make it easier for clusters of cells to cycle together than it is for lone cells to cycle in isolation.

Self-Organized Nuclear Positioning Synchronizes the Cell Cycle in Drosophila Embryos

The synchronous cleavage divisions of early embryogenesis require coordination of the cell-cycle oscillator, the dynamics of the cytoskeleton, and the cytoplasm.Yet, it remains unclear how spatially restricted biochemical signals are integrated with physical properties of the embryo to generate collective dynamics. Here, we show that synchronization of the cell cycle in Drosophila embryos requires accurate nuclear positioning, which is regulated by the cell-cycle oscillator through cortical contractility and cytoplasmic flows. We demonstrate that biochemical oscillations are initiated by local Cdk1 inactivation and spread through the activity of phosphatase PP1 to generate cortical myosin II gradients. These gradients cause cortical and cytoplasmic flows that control proper nuclear positioning. Perturbations of PP1 activity and optogenetic manipulations of cortical actomyosin disrupt nuclear spreading, resulting in loss of cell-cycle synchrony. We conclude that mitotic synchrony is established by a self-organized mechanism that integrates the cell-cycle oscillator and embryo mechanics.

Exploring the underlying mechanisms of the coupling between cell differentiation and cell cycle.

Differentiation and replication are the two major fates of the cells. They are the fundamental processes for completing the cellular functions. Although the underlying biological processes have been considerably explored for each of these processes and significant progresses have been made, global quantification and physical understanding are still challenging, especially for the relationship among them. In this study, we developed a theoretical framework for both the cell cycle and cell differentiation by exploring the associated global dynamics and their underlying relationship. We found that the dynamics of the cell cycle and cell differentiation is governed by both the landscape gradient and rotational curl flux. While landscape attracts the system down to the stable state basins, the curl flux drives the stable oscillation flow. We uncovered the irregular sombrero-shaped landscapes of the cell cycle at different developmental stages. We studied how the cells develop from undifferentiated cells to differentiated cells and how the cell cycle proceeds at different developmental stages. We investigated how the cell differentiation can influence the cell cycle where more progressive differentiation can lead to the changes of the cell cycle oscillations. In contrast, we can also quantitatively illustrate how the cell cycle can influence the cell differentiation where cell cycle regulation can lead to the changes of the differentiation processes. Through the landscape and flux analysis, we uncovered the key regulatory elements controlling the progression of the cell differentiation and cell cycle. This can help to design an effective strategy for drug discovery against associated diseases.

Heat Oscillations Driven by the Embryonic Cell Cycle Reveal the Energetic Costs of Signaling

All living systems function out of equilibrium and exchange energy in the form of heat with their environment. Thus, heat flow can inform on the energetic costs of cellular processes, which are largely unknown. Here, we have repurposed an isothermal calorimeter to measure heat flow between developing zebrafish embryos and the surrounding medium. Heat flow increased over time with cell number. Unexpectedly, a prominent oscillatory component of the heat flow, with periods matching the synchronous early reductive cleavage divisions, persisted even when DNA synthesis and mitosis were blocked by inhibitors. Instead, the heat flow oscillations were driven by the phosphorylation and dephosphorylation reactions catalyzed by the cell-cycle oscillator, the biochemical network controlling mitotic entry and exit. We propose that the high energetic cost of cell-cycle signaling reflects the significant thermodynamic burden of imposing accurate and robust timing on cell proliferation during development.

The circadian E3 ligase complex SCFFBXL3+CRY targets TLK2

We recently demonstrated that the circadian clock component CRY2 is an essential cofactor in the SCFFBXL3-mediated ubiquitination of c-MYC. Because our demonstration that CRY2 recruits phosphorylated substrates to SCFFBXL3 was unexpected, we investigated the scope of this role by searching for additional substrates of FBXL3 that require CRY1 or CRY2 as cofactors. Here, we describe an affinity purification mass spectrometry (APMS) screen through which we identified more than one hundred potential substrates of SCFFBXL3+CRY1/2, including the cell cycle regulated Tousled-like kinase, TLK2. Both CRY1 and CRY2 recruit TLK2 to SCFFBXL3, and TLK2 kinase activity is required for this interaction. Overexpression or genetic deletion of CRY1 and/or CRY2 decreases or enhances TLK2 protein abundance, respectively. These findings reinforce the idea that CRYs function as co-factors for SCFFBXL3, provide a resource of potential substrates, and establish a molecular connection between the circadian and cell cycle oscillators via CRY-modulated turnover of TLK2.

The positive circadian regulators CLOCK and BMAL1 control G2/M cell cycle transition through Cyclin B1

ABSTRACTWe previously identified a tight bidirectional phase coupling between the circadian clock and the cell cycle. To understand the role of the CLOCK/BMAL1 complex, representing the main positive regulator of the circadian oscillator, we knocked down Bmal1 or Clock in NIH3T33C mouse fibroblasts (carrying fluorescent reporters for clock and cell cycle phase) and analyzed timing of cell division in individual cells and cell populations. Inactivation of Bmal1 resulted in a loss of circadian rhythmicity and a lengthening of the cell cycle, originating from delayed G2/M transition. Subsequent molecular analysis revealed reduced levels of Cyclin B1, an important G2/M regulator, upon suppression of Bmal1 gene expression. In complete agreement with these experimental observations, simulation of Bmal1 knockdown in a computational model for coupled mammalian circadian clock and cell cycle oscillators (now incorporating Cyclin B1 induction by BMAL1) revealed a lengthening of the cell cycle. Similar data were obtained upon knockdown of Clock gene expression. In conclusion, the CLOCK/BMAL1 complex controls cell cycle progression at the level of G2/M transition through regulation of Cyclin B1 expression.

Adding phosphorylation events to the core oscillator driving the cell cycle of fission yeast.

Much is known about the regulatory elements controlling the cell cycle in fission yeast (Schizosaccharomyces pombe). This regulation is mainly done by the (cyclin-dependent kinase/cyclin) complex (Cdc2/Cdc13) that activates specific target genes and proteins via phosphorylation events during the cell cycle in a time-dependent manner. However, more work is still needed to complement the existing gaps in the current fission yeast gene regulatory network to be able to overcome abnormalities in its growth, repair and development, i.e. explain many phenomena including mitotic catastrophe. In this work we complement the previously presented core oscillator of the cell cycle of fission yeast by selected phosphorylation events and study their effects on the temporal evolution of the core oscillator based Boolean network. Thereby, we attempt to establish a regulatory link between the autonomous cell cycle oscillator and the remainder of the cell. We suggest the unclear yet regulatory effect of phosphorylation on the added components, and discuss many unreported points regarding the temporal evolution of the cell cycle and its components. To better visualize the results regardless of the programming background we developed an Android application that can be used to run the core and extended model of the fission yeast cell cycle step by step.

The Multiplex Phase Interlocker: a novel and robust molecular design synchronizing transcription and cell cycle oscillators

The Multiplex Phase Interlocker: a novel and robust molecular design synchronizing transcription and cell cycle oscillators

Non-Cell Cycle Functions of the CDK Network in Ciliogenesis: Recycling the Cell Cycle Oscillator.

Cyclin-dependent kinases are Ser/Thr protein kinases best known for their cell cycle roles, where CDK1 triggers mitotic onset in all eukaryotes. CDKs are also involved in various other cellular processes, some of which, such as transcription and centrosome duplication, are coupled to cell cycle progression. A new study suggests that the mitotic CDK network is active at low levels in non-dividing, differentiating precursors of multiciliated cells, and that it drives ciliogenesis. Manipulating the activity of CDK1 or PLK1 altered transitions between the amplification, growth, and disengagement phases, in a manner analogous to the control of passage through different phases of mitosis. How the dynamics of the mitotic kinase network are controlled in these post-mitotic cells, and whether other cell cycle regulators are also involved, remains unknown. In the present mini-review we suggest that the redeployment of cell cycle regulators to control steps of differentiation in non-dividing cells might be a more general, hitherto under-recognized, feature of cell regulation.

Delay models for the early embryonic cell cycle oscillator.

Time delays are known to play a crucial role in generating biological oscillations. The early embryonic cell cycle in the frog Xenopus laevis is one such example. Although various mathematical models of this oscillating system exist, it is not clear how to best model the required time delay. Here, we study a simple cell cycle model that produces oscillations due to the presence of an ultrasensitive, time-delayed negative feedback loop. We implement the time delay in three qualitatively different ways, using a fixed time delay, a distribution of time delays, and a delay that is state-dependent. We analyze the dynamics in all cases, and we use experimental observations to interpret our results and put constraints on unknown parameters. In doing so, we find that different implementations of the time delay can have a large impact on the resulting oscillations.