Systems biology as a foundation for genome-scale synthetic biology

As the ambitions of synthetic biology approach genome-scale engineering, comprehensive characterization of cellular systems is required, as well as a means to accurately model cell-scale molecular interactions. These requirements are coincident with the goals of systems biology and, thus, systems biology will become the foundation for genome-scale synthetic biology. Systems biology will form this foundation through its efforts to reconstruct and integrate cellular systems, develop the mathematics, theory and software tools for the accurate modeling of these integrated systems, and through evolutionary mechanisms. As genome-scale synthetic biology is so enabled, it will prove to be a positive feedback driver of systems biology by exposing and forcing researchers to confront those aspects of systems biology which are inadequately understood.     

Authentic teaching and learning through synthetic biology

Synthetic biology is an emerging engineering discipline that, if successful, will allow well-characterized biological components to be predictably and reliably built into robust organisms that achieve specific functions. Fledgling efforts to design and implement a synthetic biology curriculum for undergraduate students have shown that the co-development of this emerging discipline and its future practitioners does not undermine learning. Rather it can serve as the lynchpin of a synthetic biology curriculum. Here I describe educational goals uniquely served by synthetic biology teaching, detail ongoing curricula development efforts at MIT, and specify particular aspects of the emerging field that must develop rapidly in order to best train the next generation of synthetic biologists.     

Synthetic biology for plant genetic engineering and molecular farming

Many efforts have been put into engineering plants to improve crop yields and stress tolerance and boost the bioproduction of valuable molecules. Yet, our capabilities are still limited due to the lack of well-characterized genetic building blocks and resources for precise manipulation and given the inherently challenging properties of plant tissues. Advancements in plant synthetic biology can overcome these bottlenecks and release the full potential of engineered plants. In this review, we first discuss the recently developed plant synthetic elements from single parts to advanced circuits, software, and hardware tools expediting the engineering cycle. Next, we survey the advancements in plant biotechnology enabled by these recent resources. We conclude the review with outstanding challenges and future directions of plant synthetic biology.     

Parts plus pipes: synthetic biology approaches to metabolic engineering

Synthetic biologists combine modular biological "parts" to create higher-order devices. Metabolic engineers construct biological "pipes" by optimizing the microbial conversion of basic substrates to desired compounds. Many scientists work at the intersection of these two philosophies, employing synthetic devices to enhance metabolic engineering efforts. These integrated approaches promise to do more than simply improve product yields; they can expand the array of products that are tractable to produce biologically. In this review, we explore the application of synthetic biology techniques to next-generation metabolic engineering challenges, as well as the emerging engineering principles for biological design.     

Knowledge-making distinctions in synthetic biology

Synthetic biology is an increasingly high-profile area of research that can be understood as encompassing three broad approaches towards the synthesis of living systems: DNA-based device construction, genome-driven cell engineering and protocell creation. Each approach is characterized by different aims, methods and constructs, in addition to a range of positions on intellectual property and regulatory regimes. We identify subtle but important differences between the schools in relation to their treatments of genetic determinism, cellular context and complexity. These distinctions tie into two broader issues that define synthetic biology: the relationships between biology and engineering, and between synthesis and analysis. These themes also illuminate synthetic biology's connections to genetic and other forms of biological engineering, as well as to systems biology. We suggest that all these knowledge-making distinctions in synthetic biology raise fundamental questions about the nature of biological investigation and its relationship to the construction of biological components and systems.     

Microbial synthetic biology for human therapeutics

The emerging field of synthetic biology holds tremendous potential for developing novel drugs to treat various human conditions. The current study discusses the scope of synthetic biology for human therapeutics via microbial approach. In this context, synthetic biology aims at designing, engineering and building new microbial synthetic cells that do not pre-exist in nature as well as re-engineer existing microbes for synthesis of therapeutic products. It is expected that the construction of novel microbial genetic circuitry for human therapeutics will greatly benefit from the data generated by ??omics?? approaches and multidisciplinary nature of synthetic biology. Development of novel antimicrobial drugs and vaccines by engineering microbial systems are a promising area of research in the field of synthetic biology for human theragnostics. Expression of plant based medicinal compounds in the microbial system using synthetic biology tools is another avenue dealt in the present study. Additionally, the study suggest that the traditional medicinal knowledge can do value addition for developing novel drugs in the microbial systems using synthetic biology tools. The presented work envisions the success of synthetic biology for human therapeutics via microbial approach in a holistic manner. Keeping this in view, various legal and socio-ethical concerns emerging from the use of synthetic biology via microbial approach such as patenting, biosafety and biosecurity issues have been touched upon in the later sections.     

Bottom-up approaches in synthetic biology and biomaterials for tissue engineering applications

Synthetic biologists use engineering principles to design and construct genetic circuits for programming cells with novel functions. A bottom-up approach is commonly used to design and construct genetic circuits by piecing together functional modules that are capable of reprogramming cells with novel behavior. While genetic circuits control cell operations through the tight regulation of gene expression, a diverse array of environmental factors within the extracellular space also has a significant impact on cell behavior. This extracellular space offers an addition route for synthetic biologists to apply their engineering principles to program cell-responsive modules within the extracellular space using biomaterials. In this review, we discuss how taking a bottom-up approach to build genetic circuits using DNA modules can be applied to biomaterials for controlling cell behavior from the extracellular milieu. We suggest that, by collectively controlling intrinsic and extrinsic signals in synthetic biology and biomaterials, tissue engineering outcomes can be improved.     

The path to next generation biofuels: successes and challenges in the era of synthetic biology

Volatility of oil prices along with major concerns about climate change, oil supply security and depleting reserves have sparked renewed interest in the production of fuels from renewable resources. Recent advances in synthetic biology provide new tools for metabolic engineers to direct their strategies and construct optimal biocatalysts for the sustainable production of biofuels. Metabolic engineering and synthetic biology efforts entailing the engineering of native and de novo pathways for conversion of biomass constituents to short-chain alcohols and advanced biofuels are herewith reviewed. In the foreseeable future, formal integration of functional genomics and systems biology with synthetic biology and metabolic engineering will undoubtedly support the discovery, characterization, and engineering of new metabolic routes and more efficient microbial systems for the production of biofuels.     

Strategies for organ level tissue engineering

The field of tissue engineering has made considerable strides since it was first described in the late 1980s. The advent and subsequent boom in stem cell biology, emergence of novel technologies for biomaterial development and further understanding of developmental biology have contributed to this accelerated progress. However, continued efforts to translate tissue-engineering strategies into clinical therapies have been hampered by the problems associated with scaling up laboratory methods to produce large, complex tissues. The significant challenges faced by tissue engineers include the production of an intact vasculature within a tissue-engineered construct and recapitulation of the size and complexity of a whole organ. Here we review the basic components necessary for bioengineering organs-biomaterials, cells and bioactive molecules-and discuss various approaches for augmenting these principles to achieve organ level tissue engineering. Ultimately, the successful translation of tissue-engineered constructs into everyday clinical practice will depend upon the ability of the tissue engineer to "scale up" every aspect of the research and development process.     

Applications of cell-free protein synthesis in synthetic biology: Interfacing bio-machinery with synthetic environments

Synthetic biology is built on the synthesis, engineering, and assembly of biological parts. Proteins are the first components considered for the construction of systems with designed biological functions because proteins carry out most of the biological functions and chemical reactions inside cells. Protein synthesis is considered to comprise the most basic levels of the hierarchical structure of synthetic biology. Cell-free protein synthesis has emerged as a powerful technology that can potentially transform the concept of bioprocesses. With the ability to harness the synthetic power of biology without many of the constraints of cell-based systems, cell-free protein synthesis enables the rapid creation of protein molecules from diverse sources of genetic information. Cell-free protein synthesis is virtually free from the intrinsic constraints of cell-based methods and offers greater flexibility in system design and manipulability of biological synthetic machinery. Among its potential applications, cell-free protein synthesis can be combined with various man-made devices for rapid functional analysis of genomic sequences. This review covers recent efforts to integrate cell-free protein synthesis with various reaction devices and analytical platforms.     

Synthetic biology: a new approach to study biological pattern formation

The principles and molecular mechanisms underlying biological pattern formation are difficult to elucidate in most cases due to the overwhelming physiologic complexity associated with the natural context. The understanding of a particular mechanism, not to speak of underlying universal principles, is difficult due to the diversity and uncertainty of the biological systems. Although current genetic and biochemical approaches have greatly advanced our understanding of pattern formation, the progress mainly relies on experimental phenotypes obtained from time-consuming studies of gain or loss of function mutants. It is prevailingly considered that synthetic biology will come to the application age, but more importantly synthetic biology can be used to understand the life. Using periodic stripe pattern formation as a paradigm, we discuss how to apply synthetic biology in understanding biological pattern formation and hereafter foster the applications like tissue engineering.     

RNA synthetic biology

RNA molecules play important and diverse regulatory roles in the cell by virtue of their interaction with other nucleic acids, proteins and small molecules. Inspired by this natural versatility, researchers have engineered RNA molecules with new biological functions. In the last two years efforts in synthetic biology have produced novel, synthetic RNA components capable of regulating gene expression in vivo largely in bacteria and yeast, setting the stage for scalable and programmable cellular behavior. Immediate challenges for this emerging field include determining how computational and directed-evolution techniques can be implemented to increase the complexity of engineered RNA systems, as well as determining how such systems can be broadly extended to mammalian systems. Further challenges include designing RNA molecules to be sensors of intracellular and environmental stimuli, probes to explore the behavior of biological networks and components of engineered cellular control systems.     

TinkerCell: modular CAD tool for synthetic biology

Background

Synthetic biology brings together concepts and techniques from engineering and biology. In this field, computer-aided design (CAD) is necessary in order to bridge the gap between computational modeling and biological data. Using a CAD application, it would be possible to construct models using available biological "parts" and directly generate the DNA sequence that represents the model, thus increasing the efficiency of design and construction of synthetic networks.

Results

An application named TinkerCell has been developed in order to serve as a CAD tool for synthetic biology. TinkerCell is a visual modeling tool that supports a hierarchy of biological parts. Each part in this hierarchy consists of a set of attributes that define the part, such as sequence or rate constants. Models that are constructed using these parts can be analyzed using various third-party C and Python programs that are hosted by TinkerCell via an extensive C and Python application programming interface (API). TinkerCell supports the notion of a module, which are networks with interfaces. Such modules can be connected to each other, forming larger modular networks. TinkerCell is a free and open-source project under the Berkeley Software Distribution license. Downloads, documentation, and tutorials are available at http://www.tinkercell.com.

Conclusion

An ideal CAD application for engineering biological systems would provide features such as: building and simulating networks, analyzing robustness of networks, and searching databases for components that meet the design criteria. At the current state of synthetic biology, there are no established methods for measuring robustness or identifying components that fit a design. The same is true for databases of biological parts. TinkerCell's flexible modeling framework allows it to cope with changes in the field. Such changes may involve the way parts are characterized or the way synthetic networks are modeled and analyzed computationally. TinkerCell can readily accept third-party algorithms, allowing it to serve as a platform for testing different methods relevant to synthetic biology.     

The imminent role of protein engineering in synthetic biology

Protein engineering has for decades been a powerful tool in biotechnology for generating vast numbers of useful enzymes for industrial applications. Today, protein engineering has a crucial role in advancing the emerging field of synthetic biology, where metabolic engineering efforts alone are insufficient to maximize the full potential of synthetic biology. This article reviews the advancements in protein engineering techniques for improving biocatalytic properties to optimize engineered pathways in host systems, which are instrumental to achieve high titer production of target molecules. We also discuss the specific means by which protein engineering has improved metabolic engineering efforts and provide our assessment on its potential to continue to advance biology engineering as a whole.     

Executable cell biology

Computational modeling of biological systems is becoming increasingly important in efforts to better understand complex biological behaviors. In this review, we distinguish between two types of biological models--mathematical and computational--which differ in their representations of biological phenomena. We call the approach of constructing computational models of biological systems 'executable biology', as it focuses on the design of executable computer algorithms that mimic biological phenomena. We survey the main modeling efforts in this direction, emphasize the applicability and benefits of executable models in biological research and highlight some of the challenges that executable biology poses for biology and computer science. We claim that for executable biology to reach its full potential as a mainstream biological technique, formal and algorithmic approaches must be integrated into biological research. This will drive biology toward a more precise engineering discipline.     

Engineering microbes with synthetic biology frameworks

Typically, the outcome of biologically engineered unit operations cannot be controlled a priori due to the incorporation of ad hoc design into complex natural systems. To mitigate this problem, synthetic biology presents a systematic approach to standardizing biological components for the purpose of increasing their programmability and robustness when assembled with the aim to achieve novel biological functions. A complex engineered biological system using only standardized biological components is yet to exist. Nevertheless, current attempts to create and to implement modular, standardized biological components pave the way for the future creation of highly predictable artificial biological systems. Although synthetic biology frameworks can be applied to any biological engineering endeavor, this article will focus on providing a brief overview of advances in the field and its recent utilization for the engineering of microbes.     

A perspective of synthetic biology: assembling building blocks for novel functions

Synthetic biology is a recently emerging field that applies engineering formalisms to design and construct new biological parts, devices, and systems for novel functions or life forms that do not exist in nature. Synthetic biology relies on and shares tools from genetic engineering, bioengineering, systems biology and many other engineering disciplines. It is also different from these subjects, in both insights and approach. Applications of synthetic biology have great potential for novel contributions to established fields and for offering opportunities to answer fundamentally new biological questions. This article does not aim at a thorough survey of the literature and detailing progress in all different directions. Instead, it is intended to communicate a way of thinking for synthetic biology in which basic functional elements are defined and assembled into living systems or biomaterials with new properties and behaviors. Four major application areas with a common theme are discussed and a procedure (or "protocol") for a standard synthetic biology work is suggested.     

Synthetic biology of minimal systems

“Synthetic biology” is a concept that has developed together with, or slightly after, “systems biology”. But while systems biology aims at the full understanding of large systems by integrating more and more details into their models, synthetic biology phrases different questions, namely: what particular biological function could be obtained with a certain known subsystem of reduced complexity; can this function be manipulated or engineered in artificial environments or genetically modified organisms; and if so, how? The most prominent representation of synthetic biology has so far been microbial engineering by recombinant DNA technology, employing modular concepts known from information technology. However, there are an increasing number of biophysical groups who follow similar strategies of dissecting cellular processes and networks, trying to identify functional minimal modules that could then be combined in a bottom-up approach towards biology. These modules are so far not as particularly defined by their impact on DNA processing, but rather influenced by core fields of biophysics, such as cell mechanics and membrane dynamics. This review will give an overview of some classical and some quite new biophysical strategies for constructing minimal systems of certain cellular modules. We will show that with recent advances in understanding of cytoskeletal and membrane elements, the time might have come to experimentally challenge the concept of a minimal cell.     

Innovation at the intersection of synthetic and systems biology

The promises of modern biotechnology hinge upon the hope that we can understand microscopic cellular complexity and in doing so create novel function. In this regard, the fields of systems and synthetic biology are important for accelerating both our understanding of biological systems and our ability to quantitatively engineer cells. At the nexus of these two fields is a unique synergy that can help attain these goals. Thus, the next greatest advances in biology and biotechnology are arising at the intersection of the top-down systems approach and the bottom-up synthetic approach. Collectively, these developments enable the precise control of cellular state for systems studies and the discovery of novel parts, control strategies, and interactions for the design of robust synthetic function. This review seeks to highlight this activity as well as provide a perspective for future directions. Combining these efforts can provide novel insights into cellular function and lead to robust, novel synthetic design.     

Developmental biology and tissue engineering

Morphogenesis implies the controlled spatial organization of cells that gives rise to tissues and organs in early embryonic development. While morphogenesis is under strict genetic control, the formation of specialized biological structures of specific shape hinges on physical processes. Tissue engineering (TE) aims at reproducing morphogenesis in the laboratory, i.e., in vitro, to fabricate replacement organs for regenerative medicine. The classical approach to generate tissues/organs is by seeding and expanding cells in appropriately shaped biocompatible scaffolds, in the hope that the maturation process will result in the desired structure. To accomplish this goal more naturally and efficiently, we set up and implemented a novel TE method that is based on principles of developmental biology and employs bioprinting, the automated delivery of cellular composites into a three-dimensional (3D) biocompatible environment. The novel technology relies on the concept of tissue liquidity according to which multicellular aggregates composed of adhesive and motile cells behave in analogy with liquids: in particular, they fuse. We emphasize the major role played by tissue fusion in the embryo and explain how the parameters (surface tension, viscosity) that govern tissue fusion can be used both experimentally and theoretically to control and simulate the self-assembly of cellular spheroids into 3D living structures. The experimentally observed postprinting shape evolution of tube- and sheet-like constructs is presented. Computer simulations, based on a liquid model, support the idea that tissue liquidity may provide a mechanism for in vitro organ building.