Synthetic biology provides an opportunity for the construction and exploration of alternative solutions to biological problems - solutions different from those chosen by natural life. To this end, synthetic biologists have built new sensory systems, cellular memories, and alternative genetic codes. There is a growing interest in applying synthetic approaches to multicellular systems, especially in relation to multicellular self-organization. Here we describe a synthetic biological system that confers large-scale de novo patterning activity on 2-D and 3-D populations of mammalian cells. Instead of using the reaction-diffusion mechanisms common in real embryos, our system uses cadherin-mediated phase separation, inspired by the known phenomenon of cadherin-based sorting. An engineered self-organizing, large-scale patterning system requiring no prior spatial cue may be a significant step towards the construction of self-assembling synthetic tissues.

Optogenetic switches are emerging molecular tools for studying cellular processes as they offer higher spatiotemporal and quantitative precision than classical, chemical-based switches. Light-controllable gene expression systems designed to upregulate protein expression levels meanwhile show performances superior to their chemical-based counterparts. However, systems to reduce protein levels with similar efficiency are lagging behind. Here, we present a novel two-component, blue light-responsive optogenetic OFF switch (‘Blue-OFF’), which enables a rapid and quantitative down-regulation of a protein upon illumination. Blue-OFF combines the first light responsive repressor KRAB-EL222 with the protein degradation module B-LID (blue light-inducible degradation domain) to simultaneously control gene expression and protein stability with a single wavelength. Blue-OFF thus outperforms current optogenetic systems for controlling protein levels. The system is described by a mathematical model which aids in the choice of experimental conditions such as light intensity and illumination regime to obtain the desired outcome. This approach represents an advancement of dual-controlled optogenetic systems in which multiple photosensory modules operate synergistically. As exemplified here for the control of apoptosis in mammalian cell culture, the approach opens up novel perspectives in fundamental research and applications such as tissue engineering.

BackgroundIn mammalian development, the formation of most tissues is achieved by a relatively small repertoire of basic morphogenetic events (e.g. cell adhesion, locomotion, apoptosis, etc.), permutated in various sequences to form different tissues. Together with cell differentiation, these mechanisms allow populations of cells to organize themselves into defined geometries and structures, as simple embryos develop into complex organisms. The control of tissue morphogenesis by populations of engineered cells is a potentially very powerful but neglected aspect of synthetic biology.ResultsWe have assembled a modular library of synthetic morphogenetic driver genes to control (separately) mammalian cell adhesion, locomotion, fusion, proliferation and elective cell death. Here we describe this library and demonstrate its use in the T-REx-293 human cell line to induce each of these desired morphological behaviours on command.ConclusionsBuilding on from the simple test systems described here, we want to extend engineered control of morphogenetic cell behaviour to more complex 3D structures that can inform embryologists and may, in the future, be used in surgery and regenerative medicine, making synthetic morphology a powerful tool for developmental biology and tissue engineering.Electronic supplementary materialThe online version of this article (doi:10.1186/1754-1611-8-26) contains supplementary material, which is available to authorized users.

The objective of this study was to determine whether an antigen cocktail containing recombinantly expressed versions of most of the protective proteases of H-gal-GP, a known protective antigen from Haemonchus contortus, would confer any protection to lambs in a vaccine-challenge trial. Haemonchus contortus metalloendopeptidases, MEP1, MEP3 and MEP4, were expressed as soluble recombinant proteins in insect cells, but attempts to express the H. contortus aspartyl proteases, PEP1 and PEP2, by the same techniques were not successful. Recombinant H. contortus PEP1 was therefore expressed in Escherichia coli and refolded. Groups of sheep were immunized thrice with either native H-gal-GP, a cocktail of recombinantly expressed proteins (rMEP1, rMEP3, rMEP4 and rPep1), or adjuvant only (QuilA in PBS). All sheep were challenged with 5000 infective larvae 1 week after the final vaccination. High levels of serum antibodies that recognized H-gal-GP were detected in both the native antigen and recombinant cocktail-immunized groups by the time of challenge, but protective immunity was only observed in the group immunized with native H-gal-GP.

Bacillus thuringiensis serovar Monterrey strain BGSC 4AJ1 produced a microscopically visible capsule that reacted with a fluorescent antibody specific for the poly-gamma-d-glutamic acid (PGA) capsule of Bacillus anthracis. PGA capsule biosynthesis genes with 75%, 81%, 72%, 65% and 63% similarity, respectively, to those of the B. anthracis capBCADE cluster were present on a plasmid (pAJ1-1). Strain BGSC 4AJ1, together with five strains of Bacillus cereus that hybridized to a PGA cap gene probe, were analyzed phylogenetically using six housekeeping genes of a B. cereus multilocus sequence typing scheme. Bacillus thuringiensis BGSC 4AJ1 shared four identical alleles with B. anthracis and was the second most closely related to this bacterium of the 674 isolates in the multilocus sequence typing database. The other cap+ strains were distributed among various lineages of Clade 1 of the B. cereus group.

Classical tissue engineering is aimed mainly at producing anatomically and physiologically realistic replacements for normal human tissues. It is done either by encouraging cellular colonization of manufactured matrices or cellular recolonization of decellularized natural extracellular matrices from donor organs, or by allowing cells to self-organize into organs as they do during fetal life. For repair of normal bodies, this will be adequate but there are reasons for making unusual, non-evolved tissues (repair of unusual bodies, interface to electromechanical prostheses, incorporating living cells into life-support machines). Synthetic biology is aimed mainly at engineering cells so that they can perform custom functions: applying synthetic biological approaches to tissue engineering may be one way of engineering custom structures. In this article, we outline the ‘embryological cycle’ of patterning, differentiation and morphogenesis and review progress that has been made in constructing synthetic biological systems to reproduce these processes in new ways. The state-of-the-art remains a long way from making truly synthetic tissues, but there are now at least foundations for future work.

This study reports a proof-of-concept study as a step toward synthetic-biological morphogenesis of tissues. Events in normal animal development usually follow the sequence: patterning → differential gene expression → morphogenesis. A synthetic biological approach to development might follow a similar sequence, with each stage under the control of synthetic biological modules. The authors have constructed and published a synthetic module that drives self-organised patterning of mammalian cell populations into patches of different cell types. Here, as a proof of concept, they extend the self-patterning module with a morphogenetic effector that drives elective cell death in just one cell type. The result is a self-constructing pattern of two cell types, one of which can be selectively eliminated to leave remaining cells as a monolayer with a net-like structure. This simple device demonstrates and validates the idea of coupling synthetic biological morphogenetic effectors to synthetic biological patterning devices. It opens the path to engineering more sophisticated structures and, perhaps eventually, tissues.

Eight strains of Lactobacillus with identical partial 16S rRNA gene sequences and similar randomly amplified polymorphic DNA patterns were isolated from fermentation samples from Japanese and Scottish malt whisky distilleries. Phylogenetic analysis of almost complete 16S rRNA gene sequences from three representative strains (two from Japan, one from Scotland) placed them in the genus Lactobacillus as members of the Lactobacillus acidophilus group. Lactobacillus helveticus and Lactobacillus gallinarum were the most closely related species, with 16S rRNA gene similarities of 99?3 and 98?1 %, respectively. A similar phylogeny was derived from partial sequences of elongation factor Tu (tuf ) genes in which the alleles from the three distillery isolates were identical and shared 99?0 % similarity with L. helveticus and L. gallinarum tuf genes. S-layer (slp) gene sequences suggested different relationships among the strains and the distillery isolates no longer formed a monophyletic group. The alleles from the Japanese and Scottish strains shared only 54 % similarity. Chromosomal DNA from the distillery strains gave DNA-DNA hybridization values between 79 and 100 % but showed less than 43 and 22 % reassociation with L. helveticus and L. gallinarum DNA, respectively. The name Lactobacillus suntoryeus sp. nov. is proposed for this novel taxon; the type strain is strain SA T (=LMG 22464 T =NCIMB 14005 T ).Lactic acid bacteria comprise a natural component of the microflora of malt whisky fermentation (Simpson et al., 2001;van Beek & Priest, 2002). In well-maintained distilleries their numbers are low at the beginning of the fermentation, but, once the yeast has completed the alcoholic fermentation, they grow prolifically during the 'late lactic fermentation' and are considered to confer positive flavour notes to the spirit (Takatani & Ikemoto, 2004). The most common species encountered are Lactobacillus brevis, Lactobacillus fermentum and Lactobacillus paracasei, but numerous other Lactobacillus species, lactococci, leuconostocs and weissellas have been detected, particularly in the early stages of the fermentation, when the alcohol concentration is relatively low (van Beek & Priest, 2003).Analysis of Scotch whisky fermentations using denaturing gradient gel electrophoresis revealed an uncultured bacterium in the late stages of fermentation that was closely related to a bacterium isolated from a Japanese malt whisky fermentation and referred to as strain Y10. Phylogenetic studies based on the 16S rRNA gene sequence of strain Y10 suggested that this bacterium was a member of the Lactobacillus acidophilus group (van Beek & Priest, 2002). This group comprises L. acidophilus sensu stricto, Lactobacillus amylovorus, Lactobacillus crispatus, Lactobacillus gallinarum, Lactobacillus gasseri, Lactobacillus helveticus and Lactobacillus johnsonii (Johnson et al., 1980;Fujisawa et al., 1992;Gancheva et al., 1999), with three relatively recent additions: Lactobacillus amylolyticus, isolated from malt and beer wort (Bohak et al., 1998)...