Previous efforts to control cellular behaviour have largely relied upon various forms of genetic engineering. Once the genetic content of a living cell is modified, the behaviour of that cell typically changes as well. However, other methods of cellular control are possible. All cells sense and respond to their environment. Therefore, artificial, non-living cellular mimics could be engineered to activate or repress already existing natural sensory pathways of living cells through chemical communication. Here we describe the construction of such a system. The artificial cells expand the senses of Escherichia coli by translating a chemical message that E. coli cannot sense on its own to a molecule that activates a natural cellular response. This methodology could open new opportunities in engineering cellular behaviour without exploiting genetically modified organisms.
Artificial cells
capable of both sensing and sending chemical messages
to bacteria have yet to be built. Here we show that artificial cells
that are able to sense and synthesize quorum signaling molecules can
chemically communicate with V. fischeri, V. harveyi, E. coli, and P. aeruginosa. Activity was assessed by fluorescence, luminescence, RT-qPCR, and
RNA-seq. Two potential applications for this technology were demonstrated.
First, the extent to which artificial cells could imitate natural
cells was quantified by a type of cellular Turing test. Artificial
cells capable of sensing and in response synthesizing and releasing N-3-(oxohexanoyl)homoserine lactone showed a high degree
of likeness to natural V. fischeri under specific
test conditions. Second, artificial cells that sensed V. fischeri and in response degraded a quorum signaling molecule of P. aeruginosa (N-(3-oxododecanoyl)homoserine
lactone) were constructed, laying the foundation for future technologies
that control complex networks of natural cells.
Synthetic riboswitches can be used to control protein expression under fully defined conditions in vitro, in water-in-oil emulsions, and in vesicles. The developed system could serve as a foundation for the construction of cellular mimics that respond to molecules of our choosing.
Researchers at the National Institute for Occupational Safety and Health studied mining accidents that involved a worker entangled in, struck by, or in contact with machinery or equipment in motion. The motivation for this study came from the large number of severe accidents, i.e. accidents resulting in a fatality or permanent disability, that are occurring despite available interventions. Accident descriptions were taken from an accident database maintained by the United States Department of Labor, Mine Safety and Health Administration, and 562 accidents that occurred during 2000-2007 fit the search criteria. Machine-related accidents accounted for 41% of all severe accidents in the mining industry during this period. Machinery most often involved in these accidents included conveyors, rock bolting machines, milling machines and haulage equipment such as trucks and loaders. The most common activities associated with these accidents were operation of the machine and maintenance and repair. The current methods to safeguard workers near machinery include mechanical guarding around moving components, lockout/tagout of machine power during maintenance and backup alarms for mobile equipment. To decrease accidents further, researchers recommend additional efforts in the development of new control technologies, training materials and dissemination of information on best practices.
To facilitate the construction of cell-free genetic devices, we evaluated the ability of 17 different fluorescent proteins to give easily detectable fluorescence signals in real-time from in vitro transcription-translation reactions with a minimal system consisting of T7 RNA polymerase and E. coli translation machinery, i.e., the PUREsystem. The data were used to construct a ratiometric fluorescence assay to quantify the effect of genetic organization on in vitro expression levels. Synthetic operons with varied spacing and sequence composition between two genes that coded for fluorescent proteins were then assembled. The resulting data indicated which restriction sites and where the restriction sites should be placed in order to build genetic devices in a manner that does not interfere with protein expression. Other simple design rules were identified, such as the spacing and sequence composition influences of regions upstream and downstream of ribosome binding sites and the ability of non-AUG start codons to function in vitro.
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