Publications
15 results found
Allen ME, Hindley J, O'Toole N, et al., 2023, Biomimetic Behaviours in Hydrogel Artificial Cells through Embedded Organelles, Proceedings of the National Academy of Sciences of USA, Vol: 120, ISSN: 0027-8424
Artificial cells are biomimetic structures formed from molecular building blocks that replicate biological processes, behaviors, and architectures. Of these building blocks, hydrogels have emerged as ideal, yet underutilized candidates to provide a gel-like chassis in which to incorporate both biological and nonbiological componentry which enables the replication of cellular functionality. Here, we demonstrate a microfluidic strategy to assemble biocompatible cell-sized hydrogel-based artificial cells with a variety of different embedded functional subcompartments, which act as engineered synthetic organelles. The organelles enable the recreation of increasingly biomimetic behaviors, including stimulus-induced motility, content release through activation of membrane-associated proteins, and enzymatic communication with surrounding bioinspired compartments. In this way, we showcase a foundational strategy for the bottom–up construction of hydrogel-based artificial cell microsystems which replicate fundamental cellular behaviors, paving the way for the construction of next-generation biotechnological devices.
Gispert Contamina I, Hindley J, Pilkington C, et al., 2022, Stimuli-responsive vesicles as distributed artificial organelles for bacterial activation, Proceedings of the National Academy of Sciences of USA, Vol: 119, Pages: 1-10, ISSN: 0027-8424
Intercellular communication is a hallmark of living systems. As such, engineering artificial cells that possess this behavior has been at the heart of activities in bottom-up synthetic biology. Communication between artificial and living cells has potential to confer novel capabilities to living organisms that could be exploited in biomedicine and biotechnology. However, most current approaches rely on the exchange of chemical signals that cannot be externally controlled. Here, we report two types of remote-controlled vesicle-based artificial organelles that translate physical inputs into chemical messages that lead to bacterial activation. Upon light or temperature stimulation, artificial cell membranes are activated, releasing signaling molecules that induce protein expression in Escherichia coli. This distributed approach differs from established methods for engineering stimuli-responsive bacteria. Here, artificial cells (as opposed to bacterial cells themselves) are the design unit. Having stimuli-responsive elements compartmentalized in artificial cells has potential applications in therapeutics, tissue engineering, and bioremediation. It will underpin the design of hybrid living/nonliving systems where temporal control over population interactions can be exerted.
Allen ME, Hindley JW, Baxani DK, et al., 2022, Hydrogels as functional components in artificial cell systems, Nature Reviews Chemistry, Vol: 6, Pages: 562-578, ISSN: 2397-3358
Recent years have seen substantial efforts aimed at constructing artificial cells from various molecular components with the aim of mimicking the processes, behaviours and architectures found in biological systems. Artificial cell development ultimately aims to produce model constructs that progress our understanding of biology, as well as forming the basis for functional bio-inspired devices that can be used in fields such as therapeutic delivery, biosensing, cell therapy and bioremediation. Typically, artificial cells rely on a bilayer membrane chassis and have fluid aqueous interiors to mimic biological cells. However, a desire to more accurately replicate the gel-like properties of intracellular and extracellular biological environments has driven increasing efforts to build cell mimics based on hydrogels. This has enabled researchers to exploit some of the unique functional properties of hydrogels that have seen them deployed in fields such as tissue engineering, biomaterials and drug delivery. In this Review, we explore how hydrogels can be leveraged in the context of artificial cell development. We also discuss how hydrogels can potentially be incorporated within the next generation of artificial cells to engineer improved biological mimics and functional microsystems.
Zubaite G, Hindley JW, Ces O, et al., 2022, Dynamic reconfiguration of subcompartment architectures in artificial cells., ACS Nano, Vol: 16, ISSN: 1936-0851
Artificial cells are minimal structures constructed from biomolecular building blocks designed to mimic cellular processes, behaviors, and architectures. One near-ubiquitous feature of cellular life is the spatial organization of internal content. We know from biology that organization of content (including in membrane-bound organelles) is linked to cellular functions and that this feature is dynamic: the presence, location, and degree of compartmentalization changes over time. Vesicle-based artificial cells, however, are not currently able to mimic this fundamental cellular property. Here, we describe an artificial cell design strategy that addresses this technological bottleneck. We create a series of artificial cell architectures which possess multicompartment assemblies localized either on the inner or on the outer surface of the artificial cell membrane. Exploiting liquid-liquid phase separation, we can also engineer spatially segregated regions of condensed subcompartments attached to the cell surface, aligning with coexisting membrane domains. These structures can sense changes in environmental conditions and respond by reversibly transitioning from condensed multicompartment layers on the membrane surface to a dispersed state in the cell lumen, mimicking the dynamic compartmentalization found in biological cells. Likewise, we engineer exosome-like subcompartments that can be released to the environment. We can achieve this by using two types of triggers: chemical (addition of salts) and mechanical (by pulling membrane tethers using optical traps). These approaches allow us to control the compartmentalization state of artificial cells on population and single-cell levels.
Zhang S, Contini C, Hindley J, et al., 2021, Engineering motile aqueous phase-separated droplets via liposome stabilisation, Nature Communications, Vol: 12, Pages: 1-11, ISSN: 2041-1723
There are increasing efforts to engineer functional compartments that mimic cellular behaviours from the bottom-up. One behaviour that is receiving particular attention is motility, due to its biotechnological potential and ubiquity in living systems. Many existing platforms make use of the Marangoni effect to achieve motion in water/oil (w/o) droplet systems. However, most of these systems are unsuitable for biological applications due to biocompatibility issues caused by the presence of oil phases. Here we report a biocompatible all aqueous (w/w) PEG/dextran Pickering-like emulsion system consisting of liposome-stabilised cell-sized droplets, where the stability can be easily tuned by adjusting liposome composition and concentration. We demonstrate that the compartments are capable of negative chemotaxis: these droplets can respond to a PEG/dextran polymer gradient through directional motion down to the gradient. The biocompatibility, motility and partitioning abilities of this droplet system offers new directions to pursue research in motion-related biological processes.
Strutt R, Hindley JW, Gregg J, et al., 2021, Activating mechanosensitive channels embedded in droplet interface bilayers using membrane asymmetry, Chemical Science, Vol: 12, Pages: 2138-2145, ISSN: 2041-6520
Droplet microcompartments linked by lipid bilayers show great promise in the construction of synthetic minimal tissues. Central to controlling the flow of information in these systems are membrane proteins, which can gate in response to specific stimuli in order to control the molecular flux between membrane separated compartments. This has been demonstrated with droplet interface bilayers (DIBs) using several different membrane proteins combined with electrical, mechanical, and/or chemical activators. Here we report the activation of the bacterial mechanosensitive channel of large conductance (MscL) in a dioleoylphosphatidylcholine:dioleoylphosphatidylglycerol DIB by controlling membrane asymmetry. We show using electrical measurements that the incorporation of lysophosphatidylcholine (LPC) into one of the bilayer leaflets triggers MscL gating in a concentration-dependent manner, with partial and full activation observed at 10 and 15 mol% LPC respectively. Our findings could inspire the design of new minimal tissues where flux pathways are dynamically defined by lipid composition.
Contini C, Hindley J, Macdonald T, et al., 2020, Size dependency of gold nanoparticles interacting with model membranes, Communications Chemistry, Vol: 3, Pages: 1-12, ISSN: 2399-3669
The rapid development of nanotechnology has led to an increase in the number and variety of engineered nanomaterials in the environment. Gold nanoparticles (AuNPs) are an example of a commonly studied nanomaterial whose highly tailorable properties have generated significant interest through a wide range of research fields. In the present work, we characterise the AuNP-lipid membrane interaction by coupling qualitative data with quantitative measurements of the enthalpy change of interaction. We investigate the interactions between citrate-stabilised AuNPs ranging from 5 to 60 nm in diameter and large unilamellar vesicles acting as a model membrane system. Our results reveal the existence of two critical AuNP diameters which determine their fate when in contact with a lipid membrane. The results provide new insights into the size dependent interaction between AuNPs and lipid bilayers which is of direct relevance to nanotoxicology and to the design of NP vectors.
Zhang S, Contini C, Hindley J, et al., 2020, Engineering motile aqueous phase-separated droplets via liposome stabilisation
<jats:title>Abstract</jats:title> <jats:p>There are increasing efforts to engineer functional compartments that mimic aspects of cellular behaviour in a drive to construct an artificial cell from the bottom-up. One behaviour that is receiving particular attention is motility, due to its biotechnological potential and the fact that movement of discrete cells is a ubiquitous feature of living systems. Many existing platforms make use of the Marangoni effect to achieve motion in water/oil (w/o) droplet systems. However, most of these systems are unsuitable for biological applications due to issues with biocompatibility caused by the presence of oil phases. Here we report a biocompatible all aqueous (w/w) PEG/dextran Pickering-like emulsion system consisting of liposome-stabilized cell-sized droplets, where the stability can be easily tuned by adjusting liposome composition and concentration. We demonstrate that the compartments are capable of negative chemotaxis: if water is introduced into the emulsion system, these droplets can respond through directional motion away from PEG in the continuous phase and down to the polymer gradient with a velocity change proportional to the rearrangement of liposome stabilisers in the PEG/dextran interface. The biocompatibility, motility and partitioning abilities of this novel droplet system offers new directions to pursue research in motion-related biological processes.</jats:p>
Haylock S, Friddin M, Hindley J, et al., 2020, Membrane protein mediated bilayer communication in networks of droplet interface bilayers, Communications Chemistry, Vol: 3, ISSN: 2399-3669
Droplet interface bilayers (DIBs) are model membranes formed between lipid monolayer-encased water droplets in oil. Compared to conventional methods, one of the most unique properties of DIBs is that they can be connected together to generate multi-layered ‘tissue-like’ networks, however introducing communication pathways between these compartments typically relies on water-soluble pores that are unable to gate. Here, we show that network connectivity can instead be achieved using a water-insoluble membrane protein by successfully reconstituting a chemically activatable mutant of the mechanosensitive channel MscL into a network of DIBs. Moreover, we also show how the small molecule activator can diffuse through an open channel and across the neighbouring droplet to activate MscL present in an adjacent bilayer. This demonstration of membrane protein mediated bilayer communication could prove key toward developing the next generation of responsive bilayer networks capable of defining information flow inside a minimal tissue.
Hindley JW, Law RV, Ces O, 2020, Membrane functionalization in artificial cell engineering, SN Applied Sciences, Vol: 2, ISSN: 2523-3963
Bottom-up synthetic biology aims to construct mimics of cellular structure and behaviour known as artificial cells from a small number of molecular components. The development of this nascent field has coupled new insights in molecular biology with large translational potential for application in fields such as drug delivery and biosensing. Multiple approaches have been applied to create cell mimics, with many efforts focusing on phospholipid-based systems. This mini-review focuses on different approaches to incorporating molecular motifs as tools for lipid membrane functionalization in artificial cell construction. Such motifs range from synthetic chemical functional groups to components from extant biology that can be arranged in a ‘plug-and-play’ approach which is hard to replicate in living systems. Rationally designed artificial cells possess the promise of complex biomimetic behaviour from minimal, highly engineered chemical networks.
Hindley JW, Zheleva DG, Elani Y, et al., 2019, Building a synthetic mechanosensitive signaling pathway in compartmentalized artificial cells, Proceedings of the National Academy of Sciences, Vol: 116, Pages: 16711-16716, ISSN: 0027-8424
To date reconstitution of one of the fundamental methods of cell communication, the signaling pathway, has been unaddressed in the bottom-up construction of artificial cells (ACs). Such developments are needed to increase the functionality and biomimicry of ACs, accelerating their translation and application in biotechnology. Here we report the construction of a de novo synthetic signaling pathway in microscale nested vesicles. Vesicle cell models respond to external calcium signals through activation of an intracellular interaction between phospholipase A2 and a mechanosensitive channel present in the internal membranes, triggering content mixing between compartments and controlling cell fluorescence. Emulsion-based approaches to AC construction are therefore shown to be ideal for the quick design and testing of new signaling networks and can readily include synthetic molecules difficult to introduce to biological cells. This work represents a foundation for the engineering of multi-compartment-spanning designer pathways that can be utilised to control downstream events inside an artificial cell, leading to the assembly of micromachines capable of sensing and responding to changes in their local environment.
Karamdad K, Hindley J, Friddin MS, et al., 2018, Engineering thermoresponsive phase separated vesicles formed via emulsion phase transfer as a content-release platform, Chemical Science, Vol: 9, Pages: 4851-4858, ISSN: 2041-6520
Giant unilamellar vesicles (GUVs) are a well-established tool for the study of membrane biophysics and are increasingly used as artificial cell models and functional units in biotechnology. This trend is driven by the development of emulsion-based generation methods such as Emulsion Phase Transfer (EPT), which facilitates the encapsulation of almost any water-soluble compounds (including biomolecules) regardless of size or charge, is compatible with droplet microfluidics, and allows GUVs with asymmetric bilayers to be assembled. However, the ability to control the composition of membranes formed via EPT remains an open question; this is key as composition gives rise to an array of biophysical phenomena which can be used to add functionality to membranes. Here, we evaluate the use of GUVs constructed via this method as a platform for phase behaviour studies and take advantage of composition-dependent features to engineer thermally-responsive GUVs. For the first time, we generate ternary GUVs (DOPC/DPPC/cholesterol) using EPT, and by compensating for the lower cholesterol incorporation efficiencies, show that these possess the full range of phase behaviour displayed by electroformed GUVs. As a demonstration of the fine control afforded by this approach, we demonstrate release of dye and peptide cargo when ternary GUVs are heated through the immiscibility transition temperature, and show that release temperature can be tuned by changing vesicle composition. We show that GUVs can be individually addressed and release triggered using a laser beam. Our findings validate EPT as a suitable method for generating phase separated vesicles and provide a valuable proof-of-concept for engineering content release functionality into individually addressable vesicles, which could have a host of applications in the development of smart synthetic biosystems.
Hindley JW, Elani Y, McGilvery CM, et al., 2018, Light-triggered enzymatic reactions in nested vesicle reactors, Nature Communications, Vol: 9, Pages: 1-6, ISSN: 2041-1723
Cell-sized vesicles have tremendous potential both as miniaturised pL reaction vessels and in bottom-up synthetic biology as chassis for artificial cells. In both these areas the introduction of light-responsive modules affords increased functionality, for example, to initiate enzymatic reactions in the vesicle interior with spatiotemporal control. Here we report a system composed of nested vesicles where the inner compartments act as phototransducers, responding to ultraviolet irradiation through diacetylene polymerisation-induced pore formation to initiate enzymatic reactions. The controlled release and hydrolysis of a fluorogenic β-galactosidase substrate in the external compartment is demonstrated, where the rate of reaction can be modulated by varying ultraviolet exposure time. Such cell-like nested microreactor structures could be utilised in fields from biocatalysis through to drug delivery.
Salzmann CG, Slater B, Radaelli PG, et al., 2016, Detailed crystallographic analysis of the ice VI to ice XV hydrogen ordering phase transition, JOURNAL OF CHEMICAL PHYSICS, Vol: 145, ISSN: 0021-9606
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Messager L, Burns JR, Kim J, et al., 2016, Biomimetic hybrid nanocontainers with selective permeability, Angewandte Chemie International Edition, Vol: 55, Pages: 11106-11109, ISSN: 1521-3757
Chemistry plays a crucial role in creating synthetic analogues of biomacromolecular structures. Of particular scientific and technological interest are biomimetic vesicles that are inspired by natural membrane compartments and organelles but avoid their drawbacks, such as membrane instability and limited control over cargo transport across the boundaries. In this study, completely synthetic vesicles were developed from stable polymeric walls and easy‐to‐engineer membrane DNA nanopores. The hybrid nanocontainers feature selective permeability and permit the transport of organic molecules of 1.5 nm size. Larger enzymes (ca. 5 nm) can be encapsulated and retained within the vesicles yet remain catalytically active. The hybrid structures constitute a new type of enzymatic nanoreactor. The high tunability of the polymeric vesicles and DNA pores will be key in tailoring the nanocontainers for applications in drug delivery, bioimaging, biocatalysis, and cell mimicry.
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