Two-Way Chemical Communication between Artificial and Natural Cells

Lentini, Roberta; Martín, Noël Yeh; Forlin, Michele; Belmonte, Luca; Fontana, Jason; Cornella, Michele; Martini, Lorenzo; Tamburini, Sabrina; Bentley, William E.; Jousson, Olivier; Mansy, Sheref S.

doi:10.1021/acscentsci.6b00330

Cited by 200 publications

(241 citation statements)

References 41 publications

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“…AHL molecules are produced from acylated acyl-carrier protein (acyl-ACP) and S-adenosyl methionine (SAM) by AHL synthases such as LuxI in Vibrio fischeri ( 33). Recently, it has been shown that AHL can be produced within a CFE system by adding luxI encoding plasmid and appropriate precursors (34). Based on the idea, we tested if autolysates could produce (and sense) AHL as well.…”

Section: Resultsmentioning

confidence: 99%

Rapid and Scalable Preparation of Bacterial Lysates for Cell-Free Gene Expression

et al. 2017

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Cell-free gene expression systems are emerging as an important platform for a diverse range of synthetic biology and biotechnology applications, including production of robust field-ready biosensors. Here, we combine programmed cellular autolysis with a freeze-thaw or freeze-dry cycle to create a practical, reproducible, and a labor- and cost-effective approach for rapid production of bacterial lysates for cell-free gene expression. Using this method, robust and highly active bacterial cell lysates can be produced without specialized equipment at a wide range of scales, making cell-free gene expression easily and broadly accessible. Moreover, live autolysis strain can be freeze-dried directly and subsequently lysed upon rehydration to produce active lysate. We demonstrate the utility of autolysates for synthetic biology by regulating protein production and degradation, implementing quorum sensing, and showing quantitative protection of linear DNA templates by GamS protein. To allow versatile and sensitive β-galactosidase (LacZ) based readout we produce autolysates with no detectable background LacZ activity and use them to produce sensitive mercury(II) biosensors with LacZ-mediated colorimetric and fluorescent outputs. The autolysis approach can facilitate wider adoption of cell-free technology for cell-free gene expression as well as other synthetic biology and biotechnology applications, such as metabolic engineering, natural product biosynthesis, or proteomics.

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Section: Resultsmentioning

confidence: 99%

Rapid and Scalable Preparation of Bacterial Lysates for Cell-Free Gene Expression

et al. 2017

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“…Finally, based on recent experimental studies (26)(27)(28)(29)(30), we propose potential routes for physically realizing our QS microcapsules. A key component for QS in our system is a regulated production of chemicals.…”

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…Encapsulation of cell-free biochemical networks has been achieved using phospholipid vesicles (30) and water-in-oil microemulsion droplets (27,29). Certain small molecules used in bacterial QS, such as N-(3-oxo-hexanoyl)-L-homoserine lactone, could pass through vesicle membranes (30) and the oil phase surrounding emulsion droplets (29), thus facilitating intercellular communication. Channel proteins could be used to allow transport of other signaling species across vesicle membranes (32), and stomatocytes with large openings would also freely exchange a wide range of materials with the external fluid (33).…”

Section: Discussionmentioning

confidence: 99%

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Synthetic quorum sensing in model microcapsule colonies

Shum

Balazs

2017

Proc. Natl. Acad. Sci. U.S.A.

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Biological quorum sensing refers to the ability of cells to gauge their population density and collectively initiate a new behavior once a critical density is reached. Designing synthetic materials systems that exhibit quorum sensing-like behavior could enable the fabrication of devices with both self-recognition and selfregulating functionality. Herein, we develop models for a colony of synthetic microcapsules that communicate by producing and releasing signaling molecules. Production of the chemicals is regulated by a biomimetic negative feedback loop, the "repressilator" network. Through theory and simulation, we show that the chemical behavior of such capsules is sensitive to both the density and number of capsules in the colony. For example, decreasing the spacing between a fixed number of capsules can trigger a transition in chemical activity from the steady, repressed state to largeamplitude oscillations in chemical production. Alternatively, for a fixed density, an increase in the number of capsules in the colony can also promote a transition into the oscillatory state. This configuration-dependent behavior of the capsule colony exemplifies quorum-sensing behavior. Using our theoretical model, we predict the transitions from the steady state to oscillatory behavior as a function of the colony size and capsule density.quorum sensing | repressilator | microcapsules Q uorum sensing (QS) refers to the ability of organisms in a population to assess the number and density of individuals present, allowing a specific behavior to be initiated only when a critical threshold in the population size and density is reached. QS plays a vital role in the life cycle of bacteria (1, 2), yeast (3, 4), and slime molds (5, 6), as well as social insects (7). In microorganisms, QS is based on chemical signaling among individuals in a colony. Bacteria (1, 8), for example, produce and secrete signaling molecules, which diffuse into the surrounding medium where they can be detected by other cells in the population. Through regulatory networks, the signaling molecule acts as an autoinducer; the rate of production of the autoinducer increases with its concentration. When the cell density is low, the concentration of the signaling molecule is low, and production is maintained at a low, basal rate. The cells are considered to be in the "off" QS state. When the population density is high, each cell detects a high concentration of the signaling molecule resulting from the collective production of many nearby neighbors. The cells are switched "on," increasing production of the signaling molecule and activating further metabolic pathways that are triggered by QS. Hence, spatially separated cells interact through regulatory networks, which coordinate the collective behavior of the colony.An intriguing challenge in both synthetic biology and materials science is to design man-made systems that exhibit behavior analogous to QS, i.e., sensing and responding to the system size and density. Such synthetic QS abilities could provide a route to ...

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“…In order to achieve higher order self-organization among these artificial cell-mimicking structures, it is both desirable to separate the processes occurring in them (to create a functional identity), as well as to let them interact and communicate. Hence, communication channels based on small molecule signals have been established to mediate interactions between different types of artificial cells [4] and between artificial and bacterial cells [5] . However, membranous encapsulation typically prohibits the exchange of macromolecules as signal carriers with larger (information) channel capacity.…”

mentioning

confidence: 99%

Artificial Gel‐Based Organelles for Spatial Organization of Cell‐Free Gene Expression Reactions

Aufinger

Simmel

2018

Angew Chem Int Ed

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In biological cells, chemical processes often occur inside specialized subcellular compartments or organelles. For instance, in eukaryotes mRNA is transcribed and processed inside the nucleus, exported to the endoplasmic reticulum, and translated into the encoded protein. Inspired by this high degree of intracellular organization, we here develop gel-based artificial organelles that enable sequence-specific and programmable localization of cell-free transcription and translation reactions inside an artificial cellular system. To this end, we utilize agarose microgels covalently modified with DNA templates coding for various functions and encapsulate them into emulsion droplets. We show that RNA signals transcribed from transcription organelles can be specifically targeted to capture organelles via hybridization to the corresponding DNA addresses. We also demonstrate that mRNA molecules, produced from transcription organelles and controlled by toehold switch riboregulators, are only translated in translation organelles containing their cognate DNA triggers. Spatial confinement of transcription and translation in separate organelles is thus superficially similar to gene expression in eukaryotic cells. Combining communicating gel spheres with specialized functions opens up new possibilities for programming artificial cellular systems at the organelle level.

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Two-Way Chemical Communication between Artificial and Natural Cells

Cited by 200 publications

References 41 publications

Rapid and Scalable Preparation of Bacterial Lysates for Cell-Free Gene Expression

Rapid and Scalable Preparation of Bacterial Lysates for Cell-Free Gene Expression

Synthetic quorum sensing in model microcapsule colonies

Artificial Gel‐Based Organelles for Spatial Organization of Cell‐Free Gene Expression Reactions

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