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One promise of synthetic biology is to provide solutions for biomedical and industrial problems by rational design of added functionality in living systems. Microbes are at the forefront of this biological engineering endeavor due to their general ease of handling and their relevance in many potential applications from fermentation to therapeutics. In recent years, the field has witnessed an explosion of novel regulatory tools, from synthetic orthogonal transcription factors to posttranslational mechanisms for increased control over the behavior of synthetic circuits. Tool development has been paralleled by the discovery of principles that enable increased modularity and the management of host-circuit interactions. Engineered cell-to-cell communication bridges the scales from intracellular to population-level coordination. These developments facilitate the translation of more than a decade of circuit design into applications., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The crucial role of time-keeping has required organisms to develop sophisticated regulatory networks to ensure the reliable propagation of periodic behavior. These biological clocks have long been a focus of research; however, a clear understanding of how they maintain oscillations in the face of unpredictable environments and the inherent noise of biological systems remains elusive. Here, we review the current understanding of circadian oscillations using Drosophila melanogaster as a typical example and discuss the utility of an alternative synthetic biology approach to studying these highly intricate systems.

Although genetic mutations that alter organisms' average lifespans have been identified in aging research, our understanding of the dynamic changes during aging remains limited. Here, we integrate single-cell imaging, microfluidics, and computational modeling to investigate phenotypic divergence and cellular heterogeneity during replicative aging of single S. cerevisiae cells. Specifically, we find that isogenic cells diverge early in life toward one of two aging paths, which are characterized by distinct age-associated phenotypes. We captured the dynamics of single cells along the paths with a stochastic discrete-state model, which accurately predicts both the measured heterogeneity and the lifespan of cells on each path within a cell population. Our analysis suggests that genetic and environmental factors influence both a cell's choice of paths and the kinetics of paths themselves. Given that these factors are highly conserved throughout eukaryotes, divergent aging might represent a general scheme in cellular aging of other organisms.

Cellular aging plays an important role in many diseases, such as cancers, metabolic syndromes, and neurodegenerative disorders. There has been steady progress in identifying aging-related factors such as reactive oxygen species and genomic instability, yet an emerging challenge is to reconcile the contributions of these factors with the fact that genetically identical cells can age at significantly different rates. Such complexity requires single-cell analyses designed to unravel the interplay of aging dynamics and cell-to-cell variability. Here we use microfluidic technologies to track the replicative aging of single yeast cells and reveal that the temporal patterns of heterochromatin silencing loss regulate cellular life span. We found that cells show sporadic waves of silencing loss in the heterochromatic ribosomal DNA during the early phases of aging, followed by sustained loss of silencing preceding cell death. Isogenic cells have different lengths of the early intermittent silencing phase that largely determine their final life spans. Combining computational modeling and experimental approaches, we found that the intermittent silencing dynamics is important for longevity and is dependent on the conserved Sir2 deacetylase, whereas either sustained silencing or sustained loss of silencing shortens life span. These findings reveal that the temporal patterns of a key molecular process can directly influence cellular aging, and thus could provide guidance for the design of temporally controlled strategies to extend life span.

UnlabelledA quantitative analysis of naturally-occurring regulatory networks, especially those present in mammalian cells, is difficult due to their high complexity. Much simpler gene networks can be engineered in model organisms and analyzed as isolated regulatory modules. Recently, several synthetic networks have been constructed in mammalian systems. However, most of these engineered mammalian networks have been characterized using steady-state population level measurements. Here, we use an integrated experimental-computational approach to analyze the dynamical response of a synthetic positive feedback network in individual mammalian cells. We observe a switch-like activation of the network with variable delay times in individual cells. In agreement with a stochastic model of the network, we find that increasing the strength of the positive feedback results in a decrease in the mean delay time and a more coherent activation of individual cells. Our results are important for gaining insight into biological processes which rely on positive feedback regulation.Electronic supplementary materialThe online version of this article (doi:10.1007/s11693-009-9044-5) contains supplementary material, which is available to authorized users.

Synthetic biology enables the design of gene networks to confer specific biological functions, yet it remains a challenge to rationally engineer a biological trait as complex as longevity. A naturally occurring toggle switch underlies fate decisions toward either nucleolar or mitochondrial decline during the aging of yeast cells. We rewired this endogenous toggle to engineer an autonomous genetic clock that generates sustained oscillations between the nucleolar and mitochondrial aging processes in individual cells. These oscillations increased cellular life span through the delay of the commitment to aging that resulted from either the loss of chromatin silencing or the depletion of heme. Our results establish a connection between gene network architecture and cellular longevity that could lead to rationally designed gene circuits that slow aging. [ABSTRACT FROM AUTHOR]

The development of synthetic gene oscillators has not only demonstrated our ability toforward engineer reliable circuits in living cells, but it has also proven to be an excellent testing ground for the statistical behavior of coupled noisy oscillators. Previous experimental studies demonstrated that a shared positive feedback can reliably synchronize such oscillators, though the theoretical mechanism was not studied in detail. In the present work, we examine an experimentally motivated stochastic model for coupled degrade-and-fire gene oscillators, where a core delayed negative feedback establishes oscillations within each cell, and a shared delayed positive feedback couples all cells. We use analytic and numerical techniques to investigate conditions for one cluster and multicluster synchrony. A nonzero delay in the shared positive feedback, as expected for the experimental systems, is found to be important for synchrony to occur. [ABSTRACT FROM AUTHOR]