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Synthetic Biology
Deep Sequencing Analysis of Aptazyme Variants Based on a Pistol Ribozyme
Apr 11, 2017   ACS Synthetic Biology
Kobori S, Takahashi K, Yokobayashi Y
Deep Sequencing Analysis of Aptazyme Variants Based on a Pistol Ribozyme
Apr 11, 2017
ACS Synthetic Biology
Chemically regulated self-cleaving ribozymes, or aptazymes, are emerging as a promising class of genetic devices that allow dynamic control of gene expression in synthetic biology. However, further expansion of the limited repertoire of ribozymes and aptamers, and development of new strategies to couple the RNA elements to engineer functional aptazymes are highly desirable for synthetic biology applications. Here, we report aptazymes based on the recently identified self-cleaving pistol ribozyme class using a guanine aptamer as the molecular sensing element. Two aptazyme architectures were studied by constructing and assaying 17 728 mutants by deep sequencing. Although one of the architectures did not yield functional aptazymes, a novel aptazyme design in which the aptamer and the ribozyme were placed in tandem yielded a number of guanine-inhibited ribozymes. Detailed analysis of the extensive sequence-function data suggests a mechanism that involves a competition between two mutually exclusive RNA structures reminiscent of natural bacterial riboswitches.
CARs: Synthetic Immunoreceptors for Cancer Therapy and Beyond
Apr 18, 2017   Trends In Molecular Medicine
Chang ZL, Chen YY
CARs: Synthetic Immunoreceptors for Cancer Therapy and Beyond
Apr 18, 2017
Trends In Molecular Medicine
Chimeric antigen receptors (CARs) are versatile synthetic receptors that provide T cells with engineered specificity. Clinical success in treating B-cell malignancies has demonstrated the therapeutic potential of CAR-T cells against cancer, and efforts are underway to expand the use of engineered T cells to the treatment of diverse medical conditions, including infections and autoimmune diseases. Here, we review current understanding of the molecular properties of CARs, how this knowledge informs the rational design and characterization of novel receptors, the successes and shortcomings of CAR-T cells in the clinic, and emerging solutions for the continued improvement of CAR-T cell therapy. Copyright © 2017 Elsevier Ltd. All rights reserved.
Tunable Expression Tools Enable Single-Cell Strain Distinction in the Gut Microbiome
Apr 21, 2017   Cell
Whitaker WR, Shepherd ES, Sonnenburg JL
Tunable Expression Tools Enable Single-Cell Strain Distinction in the Gut Microbiome
Apr 21, 2017
Cell
Applying synthetic biology to engineer gut-resident microbes provides new avenues to investigate microbe-host interactions, perform diagnostics, and deliver therapeutics. Here, we describe a platform for engineering Bacteroides, the most abundant genus in the Western microbiota, which includes a process for high-throughput strain modification. We have identified a novel phage promoter and translational tuning strategy and achieved an unprecedented level of expression that enables imaging of fluorescent-protein-expressing Bacteroides stably colonizing the mouse gut. A detailed characterization of the phage promoter has provided a set of constitutive promoters that span over four logs of strength without detectable fitness burden within the gut over 14 days. These promoters function predictably over a 1,000,000-fold expression range in phylogenetically diverse Bacteroides species. With these promoters, unique fluorescent signatures were encoded to allow differentiation of six species within the gut. Fluorescent protein-based differentiation of isogenic strains revealed that priority of gut colonization determines colonic crypt occupancy. Copyright © 2017 Elsevier Inc. All rights reserved.
Engineering genetic circuit interactions within and between synthetic minimal cells
Apr 21, 2017   Nature Chemistry Add nature.com free-link Cancel
Adamala KP, Martin-Alarcon DA, Guthrie-Honea KR, Boyden ES
Engineering genetic circuit interactions within and between synthetic minimal cells
Apr 21, 2017
Nature Chemistry
Genetic circuits and reaction cascades are of great importance for synthetic biology, biochemistry and bioengineering. An open question is how to maximize the modularity of their design to enable the integration of different reaction networks and to optimize their scalability and flexibility. One option is encapsulation within liposomes, which enables chemical reactions to proceed in well-isolated environments. Here we adapt liposome encapsulation to enable the modular, controlled compartmentalization of genetic circuits and cascades. We demonstrate that it is possible to engineer genetic circuit-containing synthetic minimal cells (synells) to contain multiple-part genetic cascades, and that these cascades can be controlled by external signals as well as inter-liposomal communication without crosstalk. We also show that liposomes that contain different cascades can be fused in a controlled way so that the products of incompatible reactions can be brought together. Synells thus enable a more modular creation of synthetic biology cascades, an essential step towards their ultimate programmability.
The yeast osmostress response is carbon source dependent
Apr 21, 2017   Scientific Reports
Babazadeh R, Lahtvee PJ, Adiels CB, Goksör M, Nielsen JB, Hohmann S
The yeast osmostress response is carbon source dependent
Apr 21, 2017
Scientific Reports
Adaptation to altered osmotic conditions is a fundamental property of living cells and has been studied in detail in the yeast Saccharomyces cerevisiae. Yeast cells accumulate glycerol as compatible solute, controlled at different levels by the High Osmolarity Glycerol (HOG) response pathway. Up to now, essentially all osmostress studies in yeast have been performed with glucose as carbon and energy source, which is metabolised by glycolysis with glycerol as a by-product. Here we investigated the response of yeast to osmotic stress when yeast is respiring ethanol as carbon and energy source. Remarkably, yeast cells do not accumulate glycerol under these conditions and it appears that trehalose may partly take over the role as compatible solute. The HOG pathway is activated in very much the same way as during growth on glucose and is also required for osmotic adaptation. Slower volume recovery was observed in ethanol-grown cells as compared to glucose-grown cells. Dependence on key regulators as well as the global gene expression profile were similar in many ways to those previously observed in glucose-grown cells. However, there are indications that cells re-arrange redox-metabolism when respiration is hampered under osmostress, a feature that could not be observed in glucose-grown cells.
Leveraging microbial biosynthetic pathways for the generation of 'drop-in' biofuels
Apr 20, 2017   Current Opinion In Biotechnology
Zargar A, Bailey CB, Haushalter RW, Eiben CB, Katz L, Keasling JD
Leveraging microbial biosynthetic pathways for the generation of 'drop-in' biofuels
Apr 20, 2017
Current Opinion In Biotechnology
Advances in retooling microorganisms have enabled bioproduction of 'drop-in' biofuels, fuels that are compatible with existing spark-ignition, compression-ignition, and gas-turbine engines. As the majority of petroleum consumption in the United States consists of gasoline (47%), diesel fuel and heating oil (21%), and jet fuel (8%), 'drop-in' biofuels that replace these petrochemical sources are particularly attractive. In this review, we discuss the application of aldehyde decarbonylases to produce gasoline substitutes from fatty acid products, a recently crystallized reductase that could hydrogenate jet fuel precursors from terpene synthases, and the exquisite control of polyketide synthases to produce biofuels with desired physical properties (e.g., lower freezing points). With our increased understanding of biosynthetic logic of metabolic pathways, we discuss the unique advantages of fatty acid, terpene, and polyketide synthases for the production of bio-based gasoline, diesel and jet fuel. Copyright © 2017 Elsevier Ltd. All rights reserved.
Cover Image, Volume 114, Number 6, June 2017
Apr 20, 2017   Biotechnology And Bioengineering
Goers L, Ainsworth C, Goey CH, Kontoravdi C, Freemont PS, Polizzi KM
Cover Image, Volume 114, Number 6, June 2017
Apr 20, 2017
Biotechnology And Bioengineering
Cover Legend The cover image, by Lisa Goers et al., is based on the Article Whole-cell Escherichia coli lactate biosensor for monitoring mammalian cell cultures during biopharmaceutical production, DOI: 10.1002/bit.26254. © 2017 Wiley Periodicals, Inc.
Metabolic engineering strategies for acetoin and 2,3-butanediol production: advances and prospects
Apr 20, 2017   Critical Reviews In Biotechnology
Yang T, Rao Z, Zhang X, Xu M, Xu Z, Yang ST
Metabolic engineering strategies for acetoin and 2,3-butanediol production: advances and prospects
Apr 20, 2017
Critical Reviews In Biotechnology
Acetoin and 2,3-butanediol (2,3-BD) have a large number of industrial applications. The production of acetoin and 2,3-BD has traditionally relied on oil supplies. Microbial production of acetoin and 2,3-BD will alleviate the dependence on oil. Acetoin and 2,3-BD are neighboring metabolites in the 2,3-BD metabolic pathway of bacteria. This review summarizes metabolic engineering strategies for improvement of microbial acetoin and 2,3-BD production. We also propose enhancements to current acetoin and 2,3-BD production strategies, by offering a metabolic engineering approach that is guided by systems biology and synthetic biology.
Crystal structure of the EnvZ periplasmic domain with CHAPS
Apr 19, 2017   FEBS Letters
Hwang E, Cheong HK, Kim SY, Kwon O, Blain KY, Choe S, Yeo KJ, Jung YW, Jeon YH, Cheong C
Crystal structure of the EnvZ periplasmic domain with CHAPS
Apr 19, 2017
FEBS Letters
Bacteria sense and respond to osmolarity through the EnvZ-OmpR two-component system. The structure of the periplasmic sensor domain of EnvZ (EnvZ-PD) is not available yet. Here, we present the crystal structure of EnvZ-PD in the presence of CHAPS detergent. The structure of EnvZ-PD shows similar folding topology to the PDC domains of PhoQ, DcuS, and CitA, but distinct orientations of helices and β-hairpin structures. The CD and NMR spectra of EnvZ-PD in the presence of cholate, a major component of bile salts, are similar to those with CHAPS. Chemical cross-linking shows that the dimerization of EnvZ-PD is significantly inhibited by the CHAPS and cholate. Together with β-galactosidase assay, these results suggest that bile salts may affect the EnvZ structure and function in E. coli. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
EMMA: an Extensible Mammalian Modular Assembly toolkit for the rapid design and production of diverse expression vectors
Apr 18, 2017   ACS Synthetic Biology
Martella A, Matjusaitis M, Auxillos J, Pollard SM, Cai Y
EMMA: an Extensible Mammalian Modular Assembly toolkit for the rapid design and production of diverse expression vectors
Apr 18, 2017
ACS Synthetic Biology
Mammalian plasmid expression vectors are critical reagents underpinning many facets of research across biology, biomedical research, and the biotechnology industry. Traditional cloning methods often require laborious manual design and assembly of plasmids using tailored sequential cloning steps. This process can be protracted, complicated, expensive and error-prone. New tools and strategies that facilitate the efficient design and production of bespoke vectors would help relieve a current bottleneck for researchers. To address this, we have developed an extensible mammalian modular assembly kit (EMMA). This enables rapid and efficient modular assembly of mammalian expression vectors in a one-tube, one-step golden gate cloning reaction, using a standardized library of compatible genetic parts. The high modularity, flexibility and extensibility of EMMA provide a simple method for production of functionally diverse mammalian expression vectors. We demonstrate the value of this toolkit by constructing and validating a range of representative vectors, such as: transient and stable expression vectors (transposon based vectors), targeting vectors, inducible systems, polycistronic expression cassettes, fusion proteins, and fluorescent reporters. The method also supports simple assembly combinatorial libraries, and hierarchical assembly for production of larger multigenetic cargos. In summary, EMMA is compatible with automated production, and novel genetic parts can be easily incorporated, providing new opportunities for mammalian synthetic biology.
The E. coli molecular phenotype under different growth conditions
Apr 18, 2017   Scientific Reports
Caglar MU, Houser JR, Barnhart CS, Boutz DR, Carroll SM,   . . . . . .   , Vander Wood D, Marx CJ, Marcotte EM, Barrick JE, Wilke CO
The E. coli molecular phenotype under different growth conditions
Apr 18, 2017
Scientific Reports
Modern systems biology requires extensive, carefully curated measurements of cellular components in response to different environmental conditions. While high-throughput methods have made transcriptomics and proteomics datasets widely accessible and relatively economical to generate, systematic measurements of both mRNA and protein abundances under a wide range of different conditions are still relatively rare. Here we present a detailed, genome-wide transcriptomics and proteomics dataset of E. coli grown under 34 different conditions. Additionally, we provide measurements of doubling times and in-vivo metabolic fluxes through the central carbon metabolism. We manipulate concentrations of sodium and magnesium in the growth media, and we consider four different carbon sources glucose, gluconate, lactate, and glycerol. Moreover, samples are taken both in exponential and stationary phase, and we include two extensive time-courses, with multiple samples taken between 3 hours and 2 weeks. We find that exponential-phase samples systematically differ from stationary-phase samples, in particular at the level of mRNA. Regulatory responses to different carbon sources or salt stresses are more moderate, but we find numerous differentially expressed genes for growth on gluconate and under salt and magnesium stress. Our data set provides a rich resource for future computational modeling of E. coli gene regulation, transcription, and translation.
Light-Patterned Current Generation in a Droplet Bilayer Array
Apr 18, 2017   Scientific Reports
Restrepo Schild V, Booth MJ, Box SJ, Olof SN, Mahendran KR, Bayley H
Light-Patterned Current Generation in a Droplet Bilayer Array
Apr 18, 2017
Scientific Reports
We have created a 4 × 4 droplet bilayer array comprising light-activatable aqueous droplet bio-pixels. Aqueous droplets containing bacteriorhodopsin (bR), a light-driven proton pump, were arranged on a common hydrogel surface in lipid-containing oil. A separate lipid bilayer formed at the interface between each droplet and the hydrogel; each bilayer then incorporated bR. Electrodes in each droplet simultaneously measured the light-driven proton-pumping activities of each bio-pixel. The 4 × 4 array derived by this bottom-up synthetic biology approach can detect grey-scale images and patterns of light moving across the device, which are transduced as electrical current generated in each bio-pixel. We propose that synthetic biological light-activatable arrays, produced with soft materials, might be interfaced with living tissues to stimulate neuronal pathways.
Designing uniquely addressable bio-orthogonal synthetic scaffolds for DNA and RNA origami
Apr 17, 2017   ACS Synthetic Biology
Kozyra JW, Ceccarelli A, Torelli E, Lopiccolo A, Gu JY, Fellermann H, Stimming U, Krasnogor N
Designing uniquely addressable bio-orthogonal synthetic scaffolds for DNA and RNA origami
Apr 17, 2017
ACS Synthetic Biology
Nanotechnology and synthetic biology are rapidly converging, with DNA origami being one of the leading bridging technologies. DNA origami was shown to work well in a wide array of biotic environments. However, the large majority of extant DNA origami scaffolds utilize bacteriophages or plasmid sequences thus severely limiting its future applicability as bio-orthogonal nanotechnology platform. In this paper we present the design of biologically inert (i.e. "bio-orthogonal") origami scaffolds. The synthetic scaffolds have the additional advantage of being uniquely addressable (unlike biologically derived ones) and hence are better optimised for high-yield folding. We demonstrate our fully synthetic scaffold design with both DNA and RNA origamis and describe a protocol to produce these bio-orthogonal and uniquely addressable origami scaffolds.
Synthetic Biology: Building a custom eukaryotic genome de novo
Apr 12, 2017   Nature Reviews. Genetics
Burgess DJ
Iterative integration of multiple-copy pathway genes in Yarrowia lipolytica for heterologous β-carotene production
Apr 17, 2017   Metabolic Engineering
Gao S, Tong Y, Zhu L, Ge M, Zhang Y, Chen D, Jiang Y, Yang S
Iterative integration of multiple-copy pathway genes in Yarrowia lipolytica for heterologous β-carotene production
Apr 17, 2017
Metabolic Engineering
β-Carotene is a terpenoid molecule with high hydrophobicity that is often used as an additive in foods and feed. Previous work has demonstrated the heterologous biosynthesis of β-carotene from an intrinsic high flux of acetyl-CoA in 12 steps through 11 genes in Yarrowia lipolytica. Here, an efficient biosynthetic pathway capable of producing 100-fold more β-carotene than the baseline construct was generated using strong promoters and multiple gene copies for each of the 12 steps. Using fed-batch fermentation with an optimized medium, the engineered pathway could produce 4g/L β-carotene, which was stored in lipid droplets within engineered Y. lipolytica cells. Expansion of these cells for squalene production also demonstrated that Y. lipolytica could be an industrially relevant platform for hydrophobic terpenoid production. Copyright © 2017. Published by Elsevier Inc.
Large-scale analysis of post-translational modifications in E. coli under glucose-limiting conditions
Apr 17, 2017   BMC Genomics
Brown CW, Sridhara V, Boutz DR, Person MD, Marcotte EM, Barrick JE, Wilke CO
Large-scale analysis of post-translational modifications in E. coli under glucose-limiting conditions
Apr 17, 2017
BMC Genomics
Post-translational modification (PTM) of proteins is central to many cellular processes across all domains of life, but despite decades of study and a wealth of genomic and proteomic data the biological function of many PTMs remains unknown. This is especially true for prokaryotic PTM systems, many of which have only recently been recognized and studied in depth. It is increasingly apparent that a deep sampling of abundance across a wide range of environmental stresses, growth conditions, and PTM types, rather than simply cataloging targets for a handful of modifications, is critical to understanding the complex pathways that govern PTM deposition and downstream effects. We utilized a deeply-sampled dataset of MS/MS proteomic analysis covering 9 timepoints spanning the Escherichia coli growth cycle and an unbiased PTM search strategy to construct a temporal map of abundance for all PTMs within a 400 Da window of mass shifts. Using this map, we are able to identify novel targets and temporal patterns for N-terminal N α acetylation, C-terminal glutamylation, and asparagine deamidation. Furthermore, we identify a possible relationship between N-terminal N α acetylation and regulation of protein degradation in stationary phase, pointing to a previously unrecognized biological function for this poorly-understood PTM. Unbiased detection of PTM in MS/MS proteomics data facilitates the discovery of novel modification types and previously unobserved dynamic changes in modification across growth timepoints.
Crystal structure of RNA polymerase II from Komagataella pastoris
Apr 16, 2017   Biochemical And Biophysical Research Communications
Ehara H, Umehara T, Sekine SI, Yokoyama S
Crystal structure of RNA polymerase II from Komagataella pastoris
Apr 16, 2017
Biochemical And Biophysical Research Communications
RNA polymerase II (Pol II) is a 12-subunit protein complex that conducts the transcription of mRNA and some small RNAs. In this work, the crystal structure of Pol II from the methylotropic yeast Komagataella pastoris (Pichia pastoris) was determined. While the structure is highly homologous to that of Pol II from the budding yeast Saccharomyces cerevisiae, the stalk and clamp modules of the K. pastoris Pol II displayed large inward rotations, closing the central cleft to a greater extent than in the known S. cerevisiae Pol II structures. The conformational differences reflect the inherent flexibilities of the stalk and the clamp, as additional low-resolution structures of K. pastoris Pol II in different crystal forms revealed diverse stalk and clamp orientations. Comparisons with other eukaryotic/archaeal RNA polymerase structures in the Protein Data Bank revealed the distributions of the stalk and clamp orientations. The conformational plasticity should be essential for transcriptional functions and binding various regulatory factors. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.
Light-induced protein degradation in human-derived cells
Apr 16, 2017   Biochemical And Biophysical Research Communications
Sun W, Zhang W, Zhang C, Mao M, Zhao Y, Chen X, Yang Y
Light-induced protein degradation in human-derived cells
Apr 16, 2017
Biochemical And Biophysical Research Communications
Controlling protein degradation can be a valuable tool for posttranslational regulation of protein abundance to study complex biological systems. In the present study, we designed a light-switchable degron consisting of a light oxygen voltage (LOV) domain of Avena sativa phototropin 1 (AsLOV2) and a C-terminal degron. Our results showed that the light-switchable degron could be used for rapid and specific induction of protein degradation in HEK293 cells by light in a proteasome-dependent manner. Further studies showed that the light-switchable degron could also be utilized to mediate the degradation of secreted Gaussia princeps luciferase (GLuc), demonstrating the adaptability of the light-switchable degron in different types of protein. We suggest that the light-switchable degron offers a robust tool to control protein levels and may serves as a new and significant method for gene- and cell-based therapies. Copyright © 2017. Published by Elsevier Inc.
Concurrent haloalkanoate degradation and chlorate reduction by Pseudomonas chloritidismutans AW-1T
Apr 15, 2017   Applied And Environmental Microbiology
Peng P, Zheng Y, Koehorst JJ, Schaap PJ, Stams AJM, Smidt H, Atashgahi S
Concurrent haloalkanoate degradation and chlorate reduction by Pseudomonas chloritidismutans AW-1T
Apr 15, 2017
Applied And Environmental Microbiology
Haloalkanoates are environmental pollutants that can be degraded aerobically by microorganisms producing hydrolytic dehalogenases. However, there is lack of information about anaerobic degradation of haloalkanoates. Genome analysis of Pseudomonas chloritidismutans AW-1T, a facultative anaerobic chlorate-reducing bacterium, showed presence of two putative haloacid dehalogenase genes, l-dex and dehI, encoding an L-2-haloacid dehalogenase (L-DEX) and a halocarboxylic acid dehydrogenase (DehI). Hence, we studied concurrent degradation of haloalkanoates and chlorate as a yet unexplored trait of strain AW-1T The deduced amino acid sequences of L-DEX and DehI revealed 33-37% and 26-86% identities with biochemically/structurally characterized L-DEX and D-, DL-2-haloacid dehalogenase enzymes, respectively. Physiological experiments confirmed that strain AW-1T can grow on chloroacetate, bromoacetate and both L- and D-α-halogenated propionates with chlorate as an electron acceptor. Interestingly, growth and haloalkanoates degradation were generally faster with chlorate as an electron acceptor than with oxygen. In line with this, analyses of L-DEX and DehI dehalogenase activities using cell free extract (CFE) of strain AW-1T grown on DL-2-chloropropionate under chlorate-reducing condition showed up to 3.5-fold higher dehalogenase activity than the CFE obtained from cells grown on DL-2-chloropropionate under aerobic condition. Reverse transcription quantitative PCR showed that l-dex was expressed constitutively independent of the electron donor (haloalkanoates or acetate) or acceptor (chlorate or oxygen), whereas expression of dehI was induced by haloalkanoates. Concurrent degradation of organic and inorganic halogenated compounds by strain AW-1T represents a unique metabolic capacity in a single bacterium, providing a new piece in the puzzle of the microbial halogen cycle.IMPORTANCE Halogenated organic and inorganic compounds are important environmental pollutants that have carcinogenic and genotoxic effects on both animals and humans. Previous research studied degradation of organic and inorganic halogenated compounds separately, but not concurrently. This study shows concurrent degradation of halogenated alkanoates and chlorate as electron donor and acceptor, respectively, coupled to growth in a single bacterium, Pseudomonas chloritidismutans AW-1T Hence, besides biogenesis of molecular oxygen from chlorate reduction enabling a distinctive placement of strain AW-1T between aerobic and anaerobic microorganisms, we can now add another unique metabolic potential of this bacterium to the roster. Degradation of different halogenated compounds under anoxic conditions by a single bacterium is also of interest for the natural halogen cycle in different aquatic and terrestrial ecosystems where ample natural production of halogenated compounds has been documented. Copyright © 2017 American Society for Microbiology.
Exploring the potential of Saccharomyces cerevisiae for biopharmaceutical protein production
Apr 14, 2017   Current Opinion In Biotechnology
Wang G, Huang M, Nielsen J
Exploring the potential of Saccharomyces cerevisiae for biopharmaceutical protein production
Apr 14, 2017
Current Opinion In Biotechnology
Production of recombinant proteins by yeast plays a vital role in the biopharmaceutical industry. It is therefore desirable to develop yeast platform strains for over-production of various biopharmaceutical proteins, but this requires fundamental knowledge of the cellular machinery, especially the protein secretory pathway. Integrated analyses of multi-omics datasets can provide comprehensive understanding of cellular function, and can enable systems biology-driven and mathematical model-guided strain engineering. Rational engineering and introduction of trackable genetic modifications using synthetic biology tools, coupled with high-throughput screening are, however, also efficient approaches to relieve bottlenecks hindering high-level protein production. Here we review advances in systems biology and metabolic engineering of yeast for improving recombinant protein production. Copyright © 2017 Elsevier Ltd. All rights reserved.
Microbial response to environmental stresses: from fundamental mechanisms to practical applications
Apr 14, 2017   Applied Microbiology And Biotechnology
Guan N, Li J, Shin HD, Du G, Chen J, Liu L
Microbial response to environmental stresses: from fundamental mechanisms to practical applications
Apr 14, 2017
Applied Microbiology And Biotechnology
Environmental stresses are usually active during the process of microbial fermentation and have significant influence on microbial physiology. Microorganisms have developed a series of strategies to resist environmental stresses. For instance, they maintain the integrity and fluidity of cell membranes by modulating their structure and composition, and the permeability and activities of transporters are adjusted to control nutrient transport and ion exchange. Certain transcription factors are activated to enhance gene expression, and specific signal transduction pathways are induced to adapt to environmental changes. Besides, microbial cells also have well-established repair mechanisms that protect their macromolecules against damages inflicted by environmental stresses. Oxidative, hyperosmotic, thermal, acid, and organic solvent stresses are significant in microbial fermentation. In this review, we summarize the modus operandi by which these stresses act on cellular components, as well as the corresponding resistance mechanisms developed by microorganisms. Then, we discuss the applications of these stress resistance mechanisms on the production of industrially important chemicals. Finally, we prospect the application of systems biology and synthetic biology in the identification of resistant mechanisms and improvement of metabolic robustness of microorganisms in environmental stresses.
Synthetic associative learning in engineered multicellular consortia
Apr 13, 2017   Journal Of The Royal Society, Interface
Macia J, Vidiella B, Solé RV
Synthetic associative learning in engineered multicellular consortia
Apr 13, 2017
Journal Of The Royal Society, Interface
Associative learning (AL) is one of the key mechanisms displayed by living organisms in order to adapt to their changing environments. It was recognized early as a general trait of complex multicellular organisms but is also found in 'simpler' ones. It has also been explored within synthetic biology using molecular circuits that are directly inspired in neural network models of conditioning. These designs involve complex wiring diagrams to be implemented within one single cell, and the presence of diverse molecular wires become a challenge that might be very difficult to overcome. Here we present three alternative circuit designs based on two-cell microbial consortia able to properly display AL responses to two classes of stimuli and displaying long- and short-term memory (i.e. the association can be lost with time). These designs might be a helpful approach for engineering the human gut microbiome or even synthetic organoids, defining a new class of decision-making biological circuits capable of memory and adaptation to changing conditions. The potential implications and extensions are outlined. © 2017 The Author(s).
Transcriptomics of an extended phenotype: parasite manipulation of wasp social behaviour shifts expression of caste-related genes
Apr 13, 2017   Proceedings. Biological Sciences
Geffre AC, Liu R, Manfredini F, Beani L, Kathirithamby J, Grozinger CM, Toth AL
Transcriptomics of an extended phenotype: parasite manipulation of wasp social behaviour shifts expression of caste-related genes
Apr 13, 2017
Proceedings. Biological Sciences
Parasites can manipulate host behaviour to increase their own transmission and fitness, but the genomic mechanisms by which parasites manipulate hosts are not well understood. We investigated the relationship between the social paper wasp, Polistes dominula, and its parasite, Xenos vesparum (Insecta: Strepsiptera), to understand the effects of an obligate endoparasitoid on its host's brain transcriptome. Previous research suggests that X. vesparum shifts aspects of host social caste-related behaviour and physiology in ways that benefit the parasitoid. We hypothesized that X. vesparum-infested (stylopized) females would show a shift in caste-related brain gene expression. Specifically, we predicted that stylopized females, who would normally be workers, would show gene expression patterns resembling pre-overwintering queens (gynes), reflecting gyne-like changes in behaviour. We used RNA-sequencing data to characterize patterns of brain gene expression in stylopized females and compared these with those of unstylopized workers and gynes. In support of our hypothesis, we found that stylopized females, despite sharing numerous physiological and life-history characteristics with members of the worker caste, show gyne-shifted brain expression patterns. These data suggest that the parasitoid affects its host by exploiting phenotypic plasticity related to social caste, thus shifting naturally occurring social behaviour in a way that is beneficial to the parasitoid. © 2017 The Author(s).
A plug-and-play pathway refactoring workflow for natural product research in Escherichia coli and Saccharomyces cerevisiae
Apr 12, 2017   Biotechnology And Bioengineering
Ren H, Hu P, Zhao H
A plug-and-play pathway refactoring workflow for natural product research in Escherichia coli and Saccharomyces cerevisiae
Apr 12, 2017
Biotechnology And Bioengineering
Pathway refactoring serves as an invaluable synthetic biology tool for natural product discovery, characterization and engineering. However, the complicated and laborious molecular biology techniques largely hinder its application in natural product research, especially in a high-throughput manner. Here we report a plug-and-play pathway refactoring workflow for high-throughput, flexible pathway construction and in both Escherichia coli and Saccharomyces cerevisiae. Biosynthetic genes were firstly cloned into preassembled helper plasmids with promoters and terminators, resulting in a series of expression cassettes. These expression cassettes were further assembled using Golden Gate reaction to generate fully refactored pathways. The inclusion of spacer plasmids in this system would not only increase the flexibility for refactoring pathways with different number of genes, but also facilitate gene deletion and replacement. As a proof of concept, a total of 96 pathways for combinatorial carotenoid biosynthesis were built successfully and shown to be functional. This workflow should be generally applicable to different classes of natural products produced by various organisms. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
Engineered Regulatory Systems Modulate Gene Expression of Human Commensals in the Gut
Apr 21, 2017   Cell
Lim B, Zimmermann M, Barry NA, Goodman AL
Engineered Regulatory Systems Modulate Gene Expression of Human Commensals in the Gut
Apr 21, 2017
Cell
The gut microbiota is implicated in numerous aspects of health and disease, but dissecting these connections is challenging because genetic tools for gut anaerobes are limited. Inducible promoters are particularly valuable tools because these platforms allow real-time analysis of the contribution of microbiome gene products to community assembly, host physiology, and disease. We developed a panel of tunable expression platforms for the prominent genus Bacteroides in which gene expression is controlled by a synthetic inducer. In the absence of inducer, promoter activity is fully repressed; addition of inducer rapidly increases gene expression by four to five orders of magnitude. Because the inducer is absent in mice and their diets, Bacteroides gene expression inside the gut can be modulated by providing the inducer in drinking water. We use this system to measure the dynamic relationship between commensal sialidase activity and liberation of mucosal sialic acid, a receptor and nutrient for pathogens. VIDEO ABSTRACT. Copyright © 2017 Elsevier Inc. All rights reserved.

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