Engineering Living/Synthetic Hybrid Assemblies (LSHAs) as Functional Units for Synthetic Biology

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The overarching vision of the fellowship concerns the following question: how can biological structures (i.e. cells) be coupled with artificially-constructed elements (i.e. vesicle-based synthetic cells functionalised with molecular machinery) for the construction of living/synthetic hybrid assemblies (LSHAs)? Answering this question requires that significant technological hurdles be overcome, and this is the focus of this fellowship.Synthetic biology concerns itself with the construction of biological systems unlike those seen in nature. Synthetic cells can either be engineered from the top-down, by taking cells and manipulating their genome, or from the bottom-up, by building artificial cells from scratch using simple chemical building-blocks for user-defined functions. A technological bottleneck has meant that these two approaches have existed in isolation from one another, thus hampering the power and potential of the discipline. The LSHAs developed will serve to bridge this divide. This is of strategic importance to synthetic biology: it promises to open up a new research field and deliver applications-from biosynthesis and biosensing, to bio-chemical computing and therapeutic delivery-that would further cement the area as a key emerging technology.Strategies to construct hybrid cells will centre on using developing droplet-based microfluidics to encapsulate cells within functionalised synthetic lipid vesicles. In this way, the hybrid cells will have access to the complex biochemical pathways inherent in biological systems, yet still contain artificially-constructed elements designed from the bottom up for bespoke functionality. LSHAs can thus be considered as a novel class of 'artificial eukaryote': they will consist of a vesicle host and a cellular symbiont enjoying a mutually beneficial relationship. A set of engineering rules to biochemically interface the synthetic host with the encapsulated cells will be devised and communication routes between the two will be developed.Next, their use as a suite of cutting-edge applications via a series of proof-of-concept studies will be demonstrated. These include (i) downstream processing of secreted cellular products by chemical components contained within the vesicle, (ii) using LSHAs as a means to achieve 'membrane transplants'-the replacement of a cellular plasma membrane with an artificial membrane of defined size, composition and asymmetry, and (iii) linking up multiple LSHA units to form a large-scale network capable of processing and propagating chemical signals. This fellowship will therefore act a feasibility roadmap outlining which strategies and applications do and do not work, to aid future advances in the field.This research will be undertaken in the Department of Chemistry at Imperial College. It will fall within the bracket of the Institute of Chemical Biology, and make use of some of its state of the art microfluidic, optical trapping, and microscopy facilities specially tailored to study systems such as these. It also be conducted in partnership with Prof Booth (Kings College London), who will provide key support for the incorporation of smart, responsive behaviours via mechanosensitive pores, and with Prof Yitzhak Mastai (Bar Ilan, Israel) for the addition of chiral molecular elements into the hybrid cell.