Animal cells within a tissue typically display a striking regularity in their size. To date, the molecular mechanisms that control this uniformity are still unknown. We have previously shown that size uniformity in animal cells is promoted, in part, by size-dependent regulation of G1 length. To identify the molecular mechanisms underlying this process, we performed a large-scale small molecule screen and found that the p38 MAPK pathway is involved in coordinating cell size and cell cycle progression. Small cells display higher p38 activity and spend more time in G1 than larger cells. Inhibition of p38 MAPK leads to loss of the compensatory G1 length extension in small cells, resulting in faster proliferation, smaller cell size and increased size heterogeneity. We propose a model wherein the p38 pathway responds to changes in cell size and regulates G1 exit accordingly, to increase cell size uniformity.

Single-cell analysis is pivotal to deciphering complex phenomena like heterogeneity, bistability, and asynchronous oscillations, where a population ensemble cannot represent individual behaviors. Bulk cell-free systems, despite having unique advantages of manipulation and characterization of biochemical networks, lack the essential single-cell information to understand a class of out-of-steady-state dynamics including cell cycles. Here, by encapsulating Xenopus egg extracts in water-in-oil microemulsions, we developed artificial cells that are adjustable in sizes and periods, sustain mitotic oscillations for over 30 cycles, and function in forms from the simplest cytoplasmic-only to the more complicated ones involving nuclear dynamics, mimicking real cells. Such innate flexibility and robustness make it key to studying clock properties like tunability and stochasticity. Our results also highlight energy as an important regulator of cell cycles. We demonstrate a simple, powerful, and likely generalizable strategy of integrating strengths of single-cell approaches into conventional in vitro systems to study complex clock functions.

SUMMARY Robust biological oscillators retain the critical ability to function in the presence of environmental perturbations. Although central architectures that support robust oscillations have been extensively studied, networks containing the same core vary drastically in their potential to oscillate, and it remains elusive what peripheral modifications to the core contribute to this functional variation. Here, we have generated a complete atlas of two- and three-node oscillators computationally, then systematically analyzed the association between network structure and robustness. We found that, while certain core topologies are essential for producing a robust oscillator, local structures can substantially modulate the robustness of oscillations. Notably, local nodes receiving incoherent or coherent inputs respectively promote or attenuate the overall network robustness in an additive manner. We validated these relationships in larger-scale networks reflective of real biological oscillators. Our findings provide an explanation for why auxiliary structures not required for oscillation are evolutionarily conserved and suggest simple ways to evolve or design robust oscillators.

Background Self-sustained oscillations are a ubiquitous and vital phenomenon in living systems. From primitive single-cellular bacteria to the most sophisticated organisms, periodicities have been observed in a broad spectrum of biological processes such as neuron firing, heart beats, cell cycles, circadian rhythms, etc. Defects in these oscillators can cause diseases from insomnia to cancer. Elucidating their fundamental mechanisms is of great significance to diseases, and yet challenging, due to the complexity and diversity of these oscillators. Results Approaches in quantitative systems biology and synthetic biology have been most effective by simplifying the systems to contain only the most essential regulators. Here, we will review major progress that has been made in understanding biological oscillators using these approaches. The quantitative systems biology approach allows for identification of the essential components of an oscillator in an endogenous system. The synthetic biology approach makes use of the knowledge to design the simplest, de novo oscillators in both live cells and cell-free systems. These synthetic oscillators are tractable to further detailed analysis and manipulations. Conclusion With the recent development of biological and computational tools, both approaches have made significant achievements.

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 dropletbased 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.

High-quality guided development and environmental forcing are two routes for China’s green development, and each has different focuses and outcomes. The aim of this study is to clarify coordinated mechanisms to reveal the reasons and determine paths for China’s green development. A research framework for synergetic evolution is established, and the Haken model is applied to analyze the different effects of high-quality development that guide resource- and environmental-forcing mechanisms. This research showed that: (1) the preferred route and key factor in China’s green development is high-quality developmental guidance; (2) high-quality development and environmental-forcing mechanisms are non-coordinated, and while the former might coordinate with the latter, the latter does not; (3) mutual promotion of high-quality developmental guidance and the resource mechanism was not observed; (4) the critical point coordinated value is 0.5686 for China’s green development. Eastern China possesses a relatively high level of green development, while the north, east, west, and central areas of the country still have much progress to make.

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The proliferation-quiescence decision is a dynamic process that remains incompletely understood. Live-cell imaging with fluorescent cell cycle sensors now allows us to visualize the dynamics of cell cycle transitions and has revealed that proliferation-quiescence decisions can be highly heterogeneous, even among clonal cell lines in culture. Under normal culture conditions, cells often spontaneously enter non-cycling G0 states of varying duration and depth. This also occurs in cancer cells and G0 entry in tumors may underlie tumor dormancy and issues with cancer recurrence. Here we show that a cell cycle indicator previously shown to indicate G0 upon serum starvation, mVenus-p27K-, can also be used to monitor spontaneous quiescence in untransformed and cancer cell lines. We find that the duration of spontaneous quiescence in untransformed and cancer cells is heterogeneous and that a portion of this heterogeneity results from asynchronous proliferation-quiescence decisions in pairs of daughters after mitosis, where one daughter cell enters or remains in temporary quiescence while the other does not. We find that cancer dormancy signals influence both entry into quiescence and asynchronous proliferation-quiescence decisions after mitosis. Finally, we show that spontaneously quiescent prostate cancer cells exhibit altered expression of components of the Hippo pathway and are enriched for the stem cell markers CD133 and CD44. This suggests a hypothesis that dormancy signals could promote cancer recurrence by increasing the proportion of quiescent tumor cells poised for cell cycle re-entry with stem cell characteristics in cancer.