BBSRC-NSF/BIO - Deciphering the rules of nucleus architecture with synthetic cells and organelles

One of the hallmarks of cellular systems is the non-uniform spatial distribution of biomolecular content. Although compartmentalisation of content can be found across all life classes, it is a particularly marked feature in eukaryotes which contain a diverse array of subcellular organelles. Organelles are discrete structures within cells that exist in distinct chemical space, and are specialised to perform defined biochemical tasks. The dominant eukaryotic organelle is the nucleus which houses the genome and ensures spatial segregation of transcription and translation. It is thought that the ability of eukaryotes to compartmentalise functions in the nucleus and other organelles is behind their behavioural sophistication. Technological limitations, however, have meant that our fundamental understanding of the underpinning design principles that govern compartmentalisation in cells is lacking.

Why do cells compartmentalise content? What defines the size and number of sub-compartments? How does nuclear architecture affect DNA transcription and protein synthesis? What governs the relationship between the size of the nucleus and the size of a cell? There is a need for new experimental and modelling technologies to shed light on these questions.

Recent years have seen the emergence of bottom-up synthetic biology as a powerful new research discipline in the fundamental biosciences: re-engineering biology to decipher the rules of life by building synthetic cells. Synthetic cells are structures that mimic biological cells in form, function and behaviour. They are made by bringing together biomolecular blocks in defined combination to replicate aspects of cellular life: understanding biology by building a new biology. This research area has seen remarkable growth in recent times, partly driven by pioneering new technologies in our labs at Imperial and Caltech to model, manipulate and make new biomolecular systems.

In this collaborative and international project, our UK and US teams will work in parallel to develop new microfluidic and synthetic biology technologies to assemble synthetic cells that contain nucleus-like organelles. Together with integration with new mathematical models, we will use our synthetic cells to investigate the above unresolved fundamental questions relating to cell biology. By deconstructing and reconstructing the key cellular architectural motif of the nucleus, we aim to unravel the fundamental principles of eukaryotic architecture, and of cellular compartmentalisation more generally. The modelling and experimental platforms we will be developing will be versatile, generalisable, open access, and deskilled, allowing our enabling technologies and toolkits to be utilised by the wider academic community to tackle diverse biological challenges using synthetic cells.

Although this project will focus on questions relating to fundamental biology, the engineering rule-book we will be developing as a result of our insights, combined with the novel technologies we will be developing can be deployed for the construction of a new generation of synthetic cell devices with uses in the clinic and industry as smart therapeutic agents, as bioreactors, and in bioremediation. Moreover, the unique trans-Atlantic collaboration facilitated by this scheme will also serve as a bridge between large-scale consortia in the USA and UK dedicated to the grand challenge of building a synthetic cell from scratch.

Deciphering the rules of life is the core goal of bioscience research. Progress towards our understanding of the living world has always been reliant on new techniques, and an emerging new field - synthetic cells - now offers a new 'bottom-up' biochemical approach to ask fundamental questions difficult to answer by the traditional approaches of modification and analysis of natural cells. In 2020, the BBSRC and NSF together made synthetic cells a key priority for collaborative UK-USA research, allowing us to bring together our international team of experts to lead a synthetic cell project to understand the importance of architecture and compartmentalisation within cells. Specifically, this project will focus on the key architecture that defines eukaryotes - the nucleus - examining how compartmentalisation of DNA, transcription and regulation can optimise cellular functions and determining other fundamental properties of cells, such as their size.

Working in parallel in the UK and USA, we will first develop and optimise microfluidic and membrane engineering methods to make, load and control compartments inside synthetic cells, and develop a synergistic modelling framework that will be key to predicting and analysing synthetic nucleated cells. With these foundations in place, we will collaborate to then examine the effects of compartmentalisation on coupled biochemical processes, to investigate how transcription dynamics are affected by compartmentalisation, and to explore the rules governing the size of cells and their nuclei. These fundamental questions are well-suited for the synthetic cell approach as we can build synthetic cells with defined degrees of compartmentalisation, where the number, size and content of organelles can be controlled. We will load compartments with biochemical components to mimic the fundamentals of living cells, and then explore the parameters of cellular processes, and gain mechanistic insights on the role of subcellular architectures.