"Transient delivery of gene circuits is required in many potential applications of synthetic biology, yet the pre-steady-state processes that dominate this delivery route pose major challenges for robust circuit deployment. Here we show that site-specific recombinases can rectify undesired effects by programmable timing of gene availability in multigene circuits. We exemplify the concept with a proportional sensor for endogenous microRNA (miRNA) and show a marked reduction in its ground state leakage due to desynchronization of the circuit's repressor components and their repression target. The new sensors display a dynamic range of up to 1,000-fold compared to 20-fold in the standard configuration. We applied the approach to classify cell types on the basis of miRNA expression profile and measured >200-fold output differential between positively and negatively identified cells. We also showed major improvements in specificity with cytotoxic output. Our study opens new venues in gene circuit design via judicious temporal control of circuits' genetic makeup."
"We have taken a rational approach to redesigning the amino acid binding and aminoacyl-tRNA pairing specificities of bacterial glutaminyl-tRNA synthetase. The four-stage engineering incorporates generalizable design principles and improves the pairing efficiency of noncognate glutamate with tRNAGln by over 105-fold compared to the wild-type enzyme. Better optimized designs of the protein-RNA complex include substantial reengineering of the globular core region of the tRNA, demonstrating a role for specific tRNA nucleotides in specifying the identity of the genetically encoded amino acid. Principles emerging from this engineering effort open new prospects for combining rational and genetic selection approaches to design novel aminoacyl-tRNA synthetases that ligate noncanonical amino acids onto tRNAs. This will facilitate reconstruction of the cellular translation apparatus for applications in synthetic biology."
"While the catalog of mammalian transcripts and their expression levels in different cell types and disease states is rapidly expanding, our understanding of transcript function lags behind. We present a robust technology enabling systematic investigation of the cellular consequences of repressing or inducing individual transcripts. We identify rules for specific targeting of transcriptional repressors (CRISPRi), typically achieving 90%-99% knockdown with minimal off-target effects, and activators (CRISPRa) to endogenous genes via endonuclease-deficient Cas9. Together they enable modulation of gene expression over a ∼1,000-fold range. Using these rules, we construct genome-scale CRISPRi and CRISPRa libraries, each of which we validate with two pooled screens. Growth-based screens identify essential genes, tumor suppressors, and regulators of differentiation. Screens for sensitivity to a cholera-diphtheria toxin provide broad insights into the mechanisms of pathogen entry, retrotranslocation and toxicity. Our results establish CRISPRi and CRISPRa as powerful tools that provide rich and complementary information for mapping complex pathways."
by Pablo Cárdenas, Maaruthy Yelleswarapu, Sayane Shome, Jitendra Kumar Gupta, Eugenio Maria Battaglia, Pedro Fernandes, Alioune Ngom, and Gerd Moe-Behrens
"As we move deeper into the digital age, the social praxis of science undergoes fundamental changes, driven by new tools provided by information and communication technologies. Specifically, social networks and computing resources such as online cloud-based infrastructures and applications provide the necessary context for unprecedented innovations in modern science. These tools are leading to a planetary-scale connectivity among researchers and enable the organization of in silico research activities entirely through the cloud.
Research collaboration and management via the cloud will result in a drastic expansion of our problem-solving capacity, since groups of people with different backgrounds and expertise that openly gather around common interests are more likely to succeed at solving complex problems. Another advantage is that collaboration between individuals becomes possible regardless of their geographic location and background.
Here we present a novel, open-web application called Leukippos, which aims to apply these information and communication technologies to in silico synthetic biology projects. We describe both the underlying technology and organizational structure necessary for the platform’s operation. The synthetic biology software search engine, SynBioAppSelector, and the game, SynBrick, are examples of projects being developed on this platform."
Rice University scientists have developed a plug-and-play approach to detect interactions between proteins. Their split fluorescent proteins could greatly improve understanding of basic biological functions.
"Ability to produce embryonic stem cells will allow researchers to push faster toward cure
arvard stem cell researchers announced today that they have made a giant leap forward in the quest to find a truly effective treatment for type 1 diabetes, a disease that affects an estimated 3 million Americans at a cost of about $15 billion annually.
With human embryonic stem cells as a starting point, the scientists were for the first time able to produce, in the kind of massive quantities needed for cell transplantation and pharmaceutical purposes, human insulin-producing beta cells equivalent in most every way to normally functioning beta cells.
Doug Melton, who led the work, said he hopes to have human transplantation trials using the cells under way within a few years. Twenty-three years ago, when his infant son Sam was diagnosed with type 1 diabetes, Melton dedicated his career to finding a cure for the disease.
“We are now just one preclinical step away from the finish line,” said Melton, whose daughter Emma also has type 1 diabetes.
A report on the new work is being published today by the journal Cell."
by Lara Bereza-Malcolm , Gulay Mann , and Ashley Edwin Franks
"Whole cell microbial biosensors are offering an alternative means for rapid, on-site heavy metal detection. Based in microorganisms, biosensing constructs are designed and constructed to produce both qualitative and quantitative outputs in response to heavy metal ions. Previous microbial biosensors designs are focused on single-input constructs; however development of multiplexed systems is resulting in more flexible designs. The movement of microbial biosensors from laboratory based designs towards on-site, functioning heavy metal detectors has been hindered by the toxic nature of heavy metals, along with the lack of specificity of heavy metals promoter elements. Applying a synthetic biology approach with alternative microbial chassis may increase the robustness of microbial biosensors and mitigate these issues. Before full applications are achieved, further consideration has to be made regarding the risk and regulations of whole cell microbial biosensor use in the environment. To this end, a standard framework for future whole cell microbial biosensor design and use is proposed."
"Animal models are very important tools in drug discovery and for understanding human biology in general. However many drugs that initially show promising results in rodents fail in later stages of clinical trials. Understanding the commonalities and differences between human and rat cell signaling networks can lead to better experimental designs, improved allocation of resources and ultimately better drugs.
The sbv IMPROVER Species-Specific Network Inference challenge was designed to use the power of the crowds to build two species-specific cell signaling networks given phosphoproteomics, transcriptomics and cytokine data generated from NHBE and NRBE cells exposed to various stimuli. A common literature-inspired reference network with 220 nodes and 501 edges was also provided as prior knowledge from which challenge participants could add or remove edges but not nodes. Such a large network inference challenge not based on synthetic simulations but on real data presented unique difficulties in scoring and interpreting the results. Because any prior knowledge about the networks was already provided to the participants for reference, novel ways for scoring and aggregating the results were developed. Two human and rat consensus networks were obtained by combining all the inferred networks. Further analysis showed that major signaling pathways were conserved between the two species with only isolated components diverging, as in the case of ribosomal S6 kinase RPS6KA1. Overall, the consensus between inferred edges was relatively high with the."
"Various approaches are taken to construct synthetic cells in the laboratory, a challenging goal that became experimentally imaginable over the past two decades. The construction of protocells, which explores scenarios of the origin of life, has been the original motivations for such projects. With the advent of the synthetic biology era, bottom-up engineering approaches to synthetic cells are now conceivable. The modular design emerges as the most robust framework to construct a minimal cell from natural molecular components. Although significant advances have been made for each piece making this complex puzzle, the integration of the three fundamental parts, information–metabolism–self-organization, into cell-sized liposomes capable of sustained reproduction has failed so far. Our inability to connect these three elements is also a major limitation in this research area. New methods, such as machine learning coupled to high-throughput techniques, should be exploited to accelerate the cell-free synthesis of complex biochemical systems."