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Life
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Approaches toward Artificial Cell Construction and Applications
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Approaches toward Artificial Cell Construction and Applications

A special issue of Life (ISSN 2075-1729). This special issue belongs to the section "Synthetic Biology and Systems Biology".

Deadline for manuscript submissions: closed (30 September 2018) | Viewed by 45550

Special Issue Editor

Dr. Norikazu Ichihashi

E-Mail Website
Guest Editor
Graduate School of Frontier Biosciences, Osaka University, 1-5 Yamadaoka, Suita, Osaka 565-0871, Japan
Interests: the early evolution of life; the origin of life; artificial cell construction
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Recent advancements in biochemical techniques have enabled us to develop artificial systems with various biological functions in vitro, from basic functions, such as transcription, translation, genome replication, and lipid membrane formation, to higher-level functions, such as the expression of genes in a network, signaling through pathways, mechanisms that promote robustness, and adaptation to specific environments. One of the ultimate goals of such studies would be to construct an artificial cell. Through the construction, we can explore the boundaries between living and nonliving entities, yielding important information on the origin of life and the principles that govern it. In addition, the technologies that will need to be developed to construct artificial cells, “artificial cell technologies”, have the potential to be applied to other fields, such as evolutionary engineering, biocompatible micromachines, and new materials. This Special Issue focuses on this challenging new field.

The scope of this Special Issue encompasses all studies related to the construction of artificial cell-like systems and their parts, including replication systems, gene expression systems, metabolic systems, lipid membranes, and related cellular functions. The scope also includes chemical systems that mimic functions of biological systems. Theoretical or computational reports, reviews, and perspectives are all welcome.

I hope this Special Issue will contribute to the development of this new field.

Dr. Norikazu Ichihashi
Guest Editor

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Life is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2600 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • artificial cell
  • in vitro system
  • liposome
  • vesicle
  • replication
  • translation
  • transcription
  • metabolism
  • the origin of life
  • the principles of life
  • artificial life

Published Papers (7 papers)

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Research

Jump to: Review, Other

19 pages, 4497 KiB  
Article
Comparison between Effects of Retroactivity and Resource Competition upon Change in Downstream Reporter Genes of Synthetic Genetic Circuits
by Takefumi Moriya, Tomohiro Yamaoka, Yuki Wakayama, Shotaro Ayukawa, Zicong Zhang, Masayuki Yamamura, Shinji Wakao and Daisuke Kiga
Life 2019, 9(1), 30; https://doi.org/10.3390/life9010030 - 26 Mar 2019
Cited by 2 | Viewed by 4667
Abstract
Reporter genes have contributed to advancements in molecular biology. Binding of an upstream regulatory protein to a downstream reporter promoter allows quantification of the activity of the upstream protein produced from the corresponding gene. In studies of synthetic biology, analyses of reporter gene [...] Read more.
Reporter genes have contributed to advancements in molecular biology. Binding of an upstream regulatory protein to a downstream reporter promoter allows quantification of the activity of the upstream protein produced from the corresponding gene. In studies of synthetic biology, analyses of reporter gene activities ensure control of the cell with synthetic genetic circuits, as achieved using a combination of in silico and in vivo experiments. However, unexpected effects of downstream reporter genes on upstream regulatory genes may interfere with in vivo observations. This phenomenon is termed as retroactivity. Using in silico and in vivo experiments, we found that a different copy number of regulatory protein-binding sites in a downstream gene altered the upstream dynamics, suggesting retroactivity of reporters in this synthetic genetic oscillator. Furthermore, by separating the two sources of retroactivity (titration of the component and competition for degradation), we showed that, in the dual-feedback oscillator, the level of the fluorescent protein reporter competing for degradation with the circuits’ components is important for the stability of the oscillations. Altogether, our results indicate that the selection of reporter promoters using a combination of in silico and in vivo experiments is essential for the advanced design of genetic circuits. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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10 pages, 4693 KiB  
Article
Regeneration of Escherichia coli Giant Protoplasts to Their Original Form
by Kazuhito V. Tabata, Takao Sogo, Yoshiki Moriizumi and Hiroyuki Noji
Life 2019, 9(1), 24; https://doi.org/10.3390/life9010024 - 01 Mar 2019
Cited by 2 | Viewed by 6288
Abstract
The spheroplasts and protoplasts of cell wall-deficient (CWD) bacteria are able to revert to their original cellular morphologies through the regeneration of their cell walls. However, whether this is true for giant protoplasts (GPs), which can be as large as 10 μm in [...] Read more.
The spheroplasts and protoplasts of cell wall-deficient (CWD) bacteria are able to revert to their original cellular morphologies through the regeneration of their cell walls. However, whether this is true for giant protoplasts (GPs), which can be as large as 10 μm in diameter, is unknown. GPs can be prepared from various bacteria, including Escherichia coli and Bacillus subtilis, and also from fungi, through culture in the presence of inhibitors for cell wall synthesis or mitosis. In this report, we prepared GPs from E. coli and showed that they can return to rod-shaped bacterium, and that they are capable of colony formation. Microscopic investigation revealed that the regeneration process took place through a variety of morphological pathways. We also report the relationship between GP division and GP volume. Finally, we show that FtsZ is crucial for GP division. These results indicate that E. coli is a highly robust organism that can regenerate its original form from an irregular state, such as GP. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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14 pages, 2732 KiB  
Article
G-Protein Coupled Receptor Protein Synthesis on a Lipid Bilayer Using a Reconstituted Cell-Free Protein Synthesis System
by Belay Gessesse, Takashi Nagaike, Koji Nagata, Yoshihiro Shimizu and Takuya Ueda
Life 2018, 8(4), 54; https://doi.org/10.3390/life8040054 - 02 Nov 2018
Cited by 19 | Viewed by 7288
Abstract
Membrane proteins are important drug targets which play a pivotal role in various cellular activities. However, unlike cytosolic proteins, most of them are difficult-to-express proteins. In this study, to synthesize and produce sufficient quantities of membrane proteins for functional and structural analysis, we [...] Read more.
Membrane proteins are important drug targets which play a pivotal role in various cellular activities. However, unlike cytosolic proteins, most of them are difficult-to-express proteins. In this study, to synthesize and produce sufficient quantities of membrane proteins for functional and structural analysis, we used a bottom-up approach in a reconstituted cell-free synthesis system, the PURE system, supplemented with artificial lipid mimetics or micelles. Membrane proteins were synthesized by the cell-free system and integrated into lipid bilayers co-translationally. Membrane proteins such as the G-protein coupled receptors were expressed in the PURE system and a productivity ranging from 0.04 to 0.1 mg per mL of reaction was achieved with a correct secondary structure as predicted by circular dichroism spectrum. In addition, a ligand binding constant of 27.8 nM in lipid nanodisc and 39.4 nM in micelle was obtained by surface plasmon resonance and the membrane protein localization was confirmed by confocal microscopy in giant unilamellar vesicles. We found that our method is a promising approach to study the different classes of membrane proteins in their native-like artificial lipid bilayer environment for functional and structural studies. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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12 pages, 3083 KiB  
Article
Efficient Arrangement of the Replication Fork Trap for In Vitro Propagation of Monomeric Circular DNA in the Chromosome-Replication Cycle Reaction
by Tomonori Hasebe, Kouhei Narita, Shiomi Hidaka and Masayuki Su’etsugu
Life 2018, 8(4), 43; https://doi.org/10.3390/life8040043 - 25 Sep 2018
Cited by 12 | Viewed by 5403
Abstract
Propagation of genetic information is a fundamental prerequisite for living cells. We recently developed the replication cycle reaction (RCR), an in vitro reaction for circular DNA propagation, by reconstitution of the replication cycle of the Escherichia coli chromosome. In RCR, two replication forks [...] Read more.
Propagation of genetic information is a fundamental prerequisite for living cells. We recently developed the replication cycle reaction (RCR), an in vitro reaction for circular DNA propagation, by reconstitution of the replication cycle of the Escherichia coli chromosome. In RCR, two replication forks proceed bidirectionally from the replication origin, oriC, and meet at a region opposite oriC, yielding two copies of circular DNA. Although RCR essentially propagates supercoiled monomers, concatemer byproducts are also produced due to inefficient termination of the replication fork progression. Here, we examined the effect of the Tus-ter replication fork trap in RCR. Unexpectedly, when the fork traps were placed opposite oriC, mimicking their arrangement on the chromosome, the propagation of circular DNA was inhibited. On the other hand, fork traps flanking oriC allowed efficient propagation of circular DNA and repressed concatemer production. These findings suggest that collision of the two convergence forks through the fork trap is detrimental to repetition of the replication cycle. We further demonstrate that this detrimental effect was rescued by the UvrD helicase. These results provide insights into the way in which circular DNA monomers replicate repetitively without generating concatemers. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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11 pages, 2333 KiB  
Article
Controlled Construction of Stable Network Structure Composed of Honeycomb-Shaped Microhydrogels
by Masayuki Hayakawa, Satoshi Umeyama, Ken H. Nagai, Hiroaki Onoe and Masahiro Takinoue
Life 2018, 8(4), 38; https://doi.org/10.3390/life8040038 - 20 Sep 2018
Cited by 4 | Viewed by 4280
Abstract
Recently, the construction of models for multicellular systems such as tissues has been attracting great interest. These model systems are expected to reproduce a cell communication network and provide insight into complicated functions in living systems./Such network structures have mainly been modelled using [...] Read more.
Recently, the construction of models for multicellular systems such as tissues has been attracting great interest. These model systems are expected to reproduce a cell communication network and provide insight into complicated functions in living systems./Such network structures have mainly been modelled using a droplet and a vesicle. However, in the droplet and vesicle network, there are difficulties attributed to structural instabilities due to external stimuli and perturbations. Thus, the fabrication of a network composed of a stable component such as hydrogel is desired. In this article, the construction of a stable network composed of honeycomb-shaped microhydrogels is described. We produced the microhydrogel network using a centrifugal microfluidic technique and a photosensitive polymer. In the network, densely packed honeycomb-shaped microhydrogels were observed. Additionally, we successfully controlled the degree of packing of microhydrogels in the network by changing the centrifugal force. We believe that our stable network will contribute to the study of cell communication in multicellular systems. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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Review

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13 pages, 2245 KiB  
Review
Toward Experimental Evolution with Giant Vesicles
by Hironori Sugiyama and Taro Toyota
Life 2018, 8(4), 53; https://doi.org/10.3390/life8040053 - 31 Oct 2018
Cited by 7 | Viewed by 5436
Abstract
Experimental evolution in chemical models of cells could reveal the fundamental mechanisms of cells today. Various chemical cell models, water-in-oil emulsions, oil-on-water droplets, and vesicles have been constructed in order to conduct research on experimental evolution. In this review, firstly, recent studies with [...] Read more.
Experimental evolution in chemical models of cells could reveal the fundamental mechanisms of cells today. Various chemical cell models, water-in-oil emulsions, oil-on-water droplets, and vesicles have been constructed in order to conduct research on experimental evolution. In this review, firstly, recent studies with these candidate models are introduced and discussed with regards to the two hierarchical directions of experimental evolution (chemical evolution and evolution of a molecular self-assembly). Secondly, we suggest giant vesicles (GVs), which have diameters larger than 1 µm, as promising chemical cell models for studying experimental evolution. Thirdly, since technical difficulties still exist in conventional GV experiments, recent developments of microfluidic devices to deal with GVs are reviewed with regards to the realization of open-ended evolution in GVs. Finally, as a future perspective, we link the concept of messy chemistry to the promising, unexplored direction of experimental evolution in GVs. Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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Other

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29 pages, 4838 KiB  
Opinion
Is Research on “Synthetic Cells” Moving to the Next Level?
by Pasquale Stano
Life 2019, 9(1), 3; https://doi.org/10.3390/life9010003 - 26 Dec 2018
Cited by 56 | Viewed by 10950
Abstract
“Synthetic cells” research focuses on the construction of cell-like models by using solute-filled artificial microcompartments with a biomimetic structure. In recent years this bottom-up synthetic biology area has considerably progressed, and the field is currently experiencing a rapid expansion. Here we summarize some [...] Read more.
“Synthetic cells” research focuses on the construction of cell-like models by using solute-filled artificial microcompartments with a biomimetic structure. In recent years this bottom-up synthetic biology area has considerably progressed, and the field is currently experiencing a rapid expansion. Here we summarize some technical and theoretical aspects of synthetic cells based on gene expression and other enzymatic reactions inside liposomes, and comment on the most recent trends. Such a tour will be an occasion for asking whether times are ripe for a sort of qualitative jump toward novel SC prototypes: is research on “synthetic cells” moving to a next level? Full article
(This article belongs to the Special Issue Approaches toward Artificial Cell Construction and Applications)
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