Triggered Gene Expression in Fed‐Vesicle Microreactors with a Multifunctional Membrane

Nourian, Zohreh; Roelofsen, Wouter; Danelon, Christophe

doi:10.1002/anie.201107123

Cited by 89 publications

(78 citation statements)

References 34 publications

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“…Here, the proteins acting on the membrane are synthesized from a DNA template within liposomes, Toward the assembly of a minimal divisome 243 which radically differs from the conventional approach consisting in encapsulating purified proteins of fixed concentration. In order to feed the biosynthesis machinery with externally supplied nutrients, liposomes with enabled molecular exchange across the lipid bilayer could be prepared (Noireaux and Libchaber 2004;Nourian et al 2012). Notably, internal production of the membrane binding proteins ensures native vectorial properties, such as force generation, across the lipid bilayer, which is sometimes difficult to control when mixing purified proteins with liposomes.…”

Section: Discussionmentioning

confidence: 99%

“…We recently developed a method to trigger gene expression inside surface-tethered liposomes (Nourian et al 2012). The enzymatic apparatus of the PURE system and the DNA templates are co-encapsulated inside (sub) micrometer-sized vesicles.…”

Section: Mimicking Cell Division In Its Most Simple Formmentioning

confidence: 99%

“…We sought to express the N-BAR genes from the inside of the vesicles, following our well-established protocol (Nourian et al 2012). However, we failed observing membrane bending and filaments (not shown), presumably because the N-BAR protein cannot bind to the negatively curved membrane exposed from the liposome interior.…”

Section: Experimental Evidence Of Genetically Controlled Deformation mentioning

confidence: 99%

“…One limitation that has been invoked is the poor membrane permeability of the reaction substrates, which precludes higher yield of synthesized enzymes and catalytic products (Kuruma et al 2009). A simple mode of uptake of exogenous precursors would be to exploit the osmotic pressure-mediated transport across the lipid bilayer, which we employed to fuel the gene expression apparatus with all necessary nutrients and cofactors supplied in the external environment (Nourian et al 2012). Alternatively, ion channels or acyl transporters could be incorporated in the vesicle membrane to facilitate the diffusion of lipid precursors.…”

Section: The Lipid Biosynthesis Route For Compartment Divisionmentioning

confidence: 99%

“…Once coupled with lipid biosynthesis this mode of boundary self-reproduction holds some promise for continuous growth and division of a minimal cell. It will be important to adjust the lipid composition and annealing temperatures since gene expression in the PURE system becomes short-lived above 37°C (Nourian et al 2012).…”

Section: The Lipid Biosynthesis Route For Compartment Divisionmentioning

confidence: 99%

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Toward the assembly of a minimal divisome

2014

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The construction of an irreducible minimal cell having all essential attributes of a living system is one of the biggest challenges facing synthetic biology. One ubiquitous task accomplished by any living systems is the division of the cell envelope. Hence, the assembly of an elementary, albeit sufficient, molecular machinery that supports compartment division, is a crucial step towards the realization of self-reproducing artificial cells. Looking backward to the molecular nature of possible ancestral, supposedly more rudimentary, cell division systems may help to identify a minimal divisome. In light of a possible evolutionary pathway of division mechanisms from simple lipid vesicles toward modern life, we define two approaches for recapitulating division in primitive cells: the membrane deforming protein route and the lipid biosynthesis route. Having identified possible proteins and working mechanisms participating in membrane shape alteration, we then discuss how they could be integrated into the construction framework of a programmable minimal cell relying on gene expression inside liposomes. The protein synthesis using recombinant elements (PURE) system, a reconstituted minimal gene expression system, is conceivably the most versatile synthesis platform. As a first step towards the de novo synthesis of a divisome, we showed that the N-BAR domain protein produced from its gene could assemble onto the outer surface of liposomes and sculpt the membrane into tubular structures. We finally discuss the remaining challenges for building up a self-reproducing minimal cell, in particular the coupling of the division machinery with volume expansion and genome replication.

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Section: Discussionmentioning

confidence: 99%

Section: Mimicking Cell Division In Its Most Simple Formmentioning

confidence: 99%

Section: Experimental Evidence Of Genetically Controlled Deformation mentioning

confidence: 99%

Section: The Lipid Biosynthesis Route For Compartment Divisionmentioning

confidence: 99%

Section: The Lipid Biosynthesis Route For Compartment Divisionmentioning

confidence: 99%

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Toward the assembly of a minimal divisome

2014

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Versatile Phospholipid Assemblies for Functional Synthetic Cells and Artificial Tissues

Wang

et al. 2020

Advanced Materials

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The bottom‐up construction of a synthetic cell from nonliving building blocks capable of mimicking cellular properties and behaviors helps to understand the particular biophysical properties and working mechanisms of a cell. A synthetic cell built in this way possesses defined chemical composition and structure. Since phospholipids are native biomembrane components, their assemblies are widely used to mimic cellular structures. Here, recent developments in the formation of versatile phospholipid assemblies are described, together with the applications of these assemblies for functional membranes (protein reconstituted giant unilamellar vesicles), spherical and nonspherical protoorganelles, and functional synthetic cells, as well as the high‐order hierarchical structures of artificial tissues. Their biomedical applications are also briefly summarized. Finally, the challenges and future directions in the field of synthetic cells and artificial tissues based on phospholipid assemblies are proposed.

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Interface Engineering in Multiphase Systems toward Synthetic Cells and Organelles: From Soft Matter Fundamentals to Biomedical Applications

et al. 2020

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products of evolution over billions of years. They are featured by multiple hierarchical compartments performing specialized and exquisitely coordinated functions. The biological and genetic complexity of cells and subcellular components make understanding their physicochemical foundation piece by piece challenging. [1-5] Moreover, the mystery of how the very first cell, protocell, comes into exist in early earth scenario is unveiled yet. [6] Motivated by understanding protocell and biological cells, synthetic cells are being constructed. Started form the simplest form, aqueous droplet enclosed by a bilayer structure, researchers adopt a bottom-up approach to study machineries of biological cells, and explore possible forms of the protocell in early world conditions. [7] Synthetic cells are important to bridge the gap between cellular organism and nonliving matter. They refer to synthetic compartments that encapsulate a wide variety of molecules and mimic one or multiple cellular functions. By segregation of contents in different compartments, synthetic cells are now engineered to perform multiple processes and functions, including segregating genetic information, genetic circuits, protein synthesis, metabolism, growth, reproduce, recognition, intercellular crosstalk, and adaptivity to surroundings. [8-17] In these simplified cell models, cellular processes and signal pathways are reconstituted, providing insights for cell biology and offering new opportunities for designing smart cell-like carriers of therapeutics. [18-26] In addition, the hypothesis of "RNA world" has inspired the investigation of synthetic compartments as protocells, in which nucleotide permeation, compartmental division, the synthesis of macromolecular polynucleotide, and the polymerization of ribonucleic acids (RNA) are explored. [7,27,28] The beginning of synthesizing cell-like structures starts from amphiphiles self-assembled at liquid/liquid interfaces to form enclosed compartments. [29-32] Recently, techniques such as microfluidics facilitate the fabrication of complex compartments with high-order hierarchy with excellent control. [33-36] Formulations of compartmentalization can be diverse, there are reviews mainly focused on one particular type of synthetic compartments, such as lipid vesicles (liposomes), [22,30,37,38] polymer vesicles (polymersomes), [39-43] lipid-polymer hybrid Synthetic cells have a major role in gaining insight into the complex biological processes of living cells; they also give rise to a range of emerging applications from gene delivery to enzymatic nanoreactors. Living cells rely on compartmentalization to orchestrate reaction networks for specialized and coordinated functions. Principally, the compartmentalization has been an essential engineering theme in constructing cell-mimicking systems. Here, efforts to engineer liquid-liquid interfaces of multiphase systems into membrane-bounded and membraneless compartments, which include lipid vesicles, polymer vesicles, colloidosomes, hybrids, and coacervate droplets,...

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Triggered Gene Expression in Fed‐Vesicle Microreactors with a Multifunctional Membrane

Cited by 89 publications

References 34 publications

Toward the assembly of a minimal divisome

Toward the assembly of a minimal divisome

Versatile Phospholipid Assemblies for Functional Synthetic Cells and Artificial Tissues

Interface Engineering in Multiphase Systems toward Synthetic Cells and Organelles: From Soft Matter Fundamentals to Biomedical Applications

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