Mammalian synthetic biology: engineering of sophisticated gene networks

J Biotechnol. 2007 Jul 15;130(4):329-45. Epub 2007 May 24.

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Abstract:

With the recent development of a wide range of inducible mammalian transgene control systems it has now become possible to create functional synthetic gene networks by linking and connecting systems into various configurations. The past 5 years has thus seen the design and construction of the first synthetic mammalian gene regulatory networks. These networks have built upon pioneering advances in prokaryotic synthetic networks and possess an impressive range of functionalities that will some day enable the engineering of sophisticated inter- and intra-cellular functions to become a reality. At a relatively simple level, the modular linking of transcriptional components has enabled the creation of genetic networks that are strongly analogous to the architectural design and functionality of electronic circuits. Thus, by combining components in different serial or parallel configurations it is possible to produce networks that follow strict logic in integrating multiple independent signals (logic gates and transcriptional cascades) or which temporally modify input signals (time-delay circuits). Progressing in terms of sophistication, synthetic transcriptional networks have also been constructed which emulate naturally occurring genetic properties, such as bistability or dynamic instability. Toggle switches which possess "memory" so as to remember transient administered inputs, hysteric switches which are resistant to stochastic fluctuations in inputs, and oscillatory networks which produce regularly timed expression outputs, are all examples of networks that have been constructed using such properties. Initial steps have also been made in designing the above networks to respond not only to exogenous signals, but also endogenous signals that may be associated with aberrant cellular function or physiology thereby providing a means for tightly controlled gene therapy applications. Moving beyond pure transcriptional control, synthetic networks have also been created which utilize phenomena, such as post-transcriptional silencing, translational control, or inter-cellular signaling to produce novel network-based control both within and between cells. It is envisaged in the not-too-distant future that these networks will provide the basis for highly sophisticated genetic manipulations in biopharmaceutical manufacturing, gene therapy and tissue engineering applications.

Programmable cells: Interfacing natural and engineered gene networks

Article From Proceedings of the National Academy of Sciences of the United States of America.

(Full-text available)

Abstract:

Novel cellular behaviors and characteristics can be obtained by coupling engineered gene networks to the cell's natural regulatory circuitry through appropriately designed input and output interfaces. Here, we demonstrate how an engineered genetic circuit can be used to construct cells that respond to biological signals in a predetermined and programmable fashion. We employ a modular design strategy to create Escherichia coli strains where a genetic toggle switch is interfaced with: (i) the SOS signaling pathway responding to DNA damage, and (ii) a transgenic quorum sensing signaling pathway from Vibrio fischeri. The genetic toggle switch endows these strains with binary response dynamics and an epigenetic inheritance that supports a persistent phenotypic alteration in response to transient signals. These features are exploited to engineer cells that form biofilms in response to DNA-damaging agents and cells that activate protein synthesis when the cell population reaches a critical density. Our work represents a step toward the development of “plug-and-play” genetic circuitry that can be used to create cells with programmable behaviors.

Here is a fine discussion of the above article from an Openwetware blog.

Imaging brain electric signals with genetically targeted voltage-sensitive fluorescent proteins

Nature Methods

 

7,

 

643–649

 

(2010)

 

doi:10.1038/nmeth.1479

Received

 

23 December 2009

 

Accepted

 

07 June 2010

 

Published online

 

11 July 2010

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Abstract:
Cortical information processing relies on synaptic interactions between diverse classes of neurons with distinct electrophysiological and connection properties. Uncovering the operational principles of these elaborate circuits requires the probing of electrical activity from selected populations of defined neurons. Here we show that genetically encoded voltage-sensitive fluorescent proteins (VSFPs) provide an optical voltage report from targeted neurons in culture, acute brain slices and living mice. By expressing VSFPs in pyramidal cells of mouse somatosensory cortex, we also demonstrate that these probes can report cortical electrical responses to single sensory stimuli in vivo. These protein-based voltage probes will facilitate the analysis of cortical circuits in genetically defined cell populations and are hence a valuable addition to the optogenetic toolbox.

Clarifying brain structure, literally

Nature Methods

 

8,

 

793

 

(2011)

 

doi:10.1038/nmeth.1720

Published online

 

29 September 2011

A fluorescence-compatible tissue-clearing reagent enables light microscopy–based imaging deep in the mouse brain.

In The Invisible Man, a science fiction novella by Herbert G. Wells, the protagonist is a scientist who finds a way to make the human body invisible by changing its refractive index to prevent the bending and reflection of light. In a recent report, Atsushi Miyawaki and his colleagues at RIKEN described the development of a tissue-clearing reagent with similar effects, bridging the gap between science and fiction and enabling fluorescence-based imaging of biological tissues at unprecedented depth and subcellular resolution.

High-resolution microscopy methods and fluorescence-based labeling techniques have enabled the three-dimensional imaging and reconstruction of defined cellular populations in a variety of biological tissues. However, axial resolution and imaging depth are often limited by the intrinsic opacity of biological specimens. For example, in visualizing the mammalian brain, light microscopy–based advances have been confined to the few hundred micrometers under the organ's surface. Alternatively, mechanical sectioning or insertion of minuscule endoscopes can be used to access deeper structures, but such approaches are inevitably laborious, invasive or of limited perspective.

A GFP for RNA

Researchers describe a GFP mimic for fluorescently labeling RNA molecules.

The genetically encodable protein tag GFP and the rainbow of fluorescent variants it inspired have been indispensible for cell biology. Tagging RNAs in cells, however, is not so straightforward.

Samie Jaffrey's lab at Weill Medical College of Cornell University has long been interested in studying the role of RNAs in axon guidance. However, Jaffrey was frustrated that simple tools for visualizing RNAs were not available.

(This article relates to me, as well, because my senior seminar was on axon guidance, and I did not cover RNA mechanisms at all because I did not find any.)