Developing a synthetic signalling system capable of the precise spatial and temporal control of protein function in living cells

The aim of synthetic biology is to modify cells or biological systems in ways that produce societal and economic benefits in the areas of healthcare, energy production, food security and the environment. Eukaryotic cells hold huge potential in synthetic biology, however, due to their complexity, modifying them using conventional synthetic biology approaches, which involve constructing gene networks, is challenging. In contrast, directly regulating the function of proteins inside these cells may offer a simpler method to control their behaviour. A number of options, including antibodies, are currently available to do this, but the drawback of these approaches is that they cannot be easily switched off, so they affect the cell's behaviour continuously. In order to control cell behaviour more precisely, we need to develop a new system that can be turned on or off rapidly. The Ideal system would comprise a switch-like 'effector' protein inside the cell that can be engineered to bind and regulate other proteins, and a 'receptor' protein on the cell surface that can switch the effector protein on when a specific chemical is added to its environment. G protein-coupled receptor (GPCR) signalling pathways are natural systems that cells use to communicate; they are therefore ideal templates for the development of a new tool to control cell behaviour. GPCRs are cell-surface receptors that detect chemicals outside the cell, and activate effector proteins, called 'G proteins', inside the cell. The G proteins act as switches, which, when activated by GPCRs, bind and regulate their target protein, before switching themselves off in a time-dependent manner. Until now, it has not been possible to modify G proteins to bind and regulate different cellular proteins because of the high degree of complexity that has evolved within these pathways. However, I recently developed a simplified G protein that may, for the first time, make this possible.

This proposal aims to modify the GPCR signalling pathway to create a novel cell-based tool that will allow us to control the activity of different proteins inside live eukaryotic cells. This will enable us to either study the protein's function in real time or directly control cellular processes and behaviours. The key component of this system will be the simplified G protein, which will be modified so that it can bind and regulate different cellular proteins. This G protein can be activated by either native GPCRs, in which case the tool could be controlled using naturally occurring chemicals (such as a hormone), or a modified GPCR, in which case it could be controlled by a specially designed chemical. This tool will have applications in several different areas. First, it will underpin basic research to understand the function of native cellular proteins that are central to the health and disease of animals and humans. Second, it will have applications in the development of a range of cell-based biosensors capable of detecting hundreds of physiologically-relevant chemicals in real time. Third, it will facilitate research in the field of regenerative medicine, by facilitating the precise control of human cell behaviour, for example, cell division. The first four years of the Fellowship will be used to develop and optimise the tool, and to foster partnerships with companies that are capable of translating it into viable healthcare products. The final three years will focus on both developing the biosensor applications, with the aim of producing miniaturised medical diagnostic devices, and exploring applications to control human cell division, with the aim of developing improved treatments for degenerative conditions such as osteoarthritis. The long-term implication of this research will be a novel cell-based tool that will benefit both academic and industrial researchers, by simplifying the implementation of synthetic biology in eukaryotic cells and expediting its promised societal and economic benefits.

The output of this proposal will be a novel cell-based tool capable of the precise control of protein activity in live eukaryotic cells. It will have applications in studying the function, localisation and dynamics of proteins in yeast, animal and human cells, and will enable the direct regulation of cellular processes and behaviours in these organisms. This research will have a wide range of potentially applications in both fundamental and applied research covering multiple bioscience disciplines, including synthetic biology (synbio), biotechnology, cell biology and regenerative medicine. It will directly benefit researchers in both academia and industry, as well as those in government- or charitably-funded institutions. Team members will develop skills in state-of-the-art experimental and computational methods, which will be transferable to careers in academic, public sector, charitable and industrial environments.

The short-term (<5 years), medium-term (5-10 years) and long-term (>10 years) implications of this research are summarised below. First, the tool can be used to study the function, localisation and dynamics of intracellular proteins that are central to the health and disease of animals and humans. In the short-term this will benefit both academic and industrial researchers studying a wide variety of intracellular processes, by enabling them to inhibit the function of key cellular proteins in real time. In the long term this will result in a more complete understanding of numerous physiological or pathophysiological states and should lead to the development of novel therapeutics or disease prevention strategies. Second, it can be used to develop a range of novel biosensors that can detect hundreds of different human hormones, metabolites and disease markers in real time. In the medium term this will lead to a range of industrial applications, including, the development of next-generation wearable medical sensors. Third, it can be used to directly regulate cellular processes and behaviours, for example, cell division. In the short term, this will benefit academic and industrial researchers in the field of tissue engineering, by facilitating the large-scale production of primary human cells. In the long term it will benefit regenerative medicine companies by enabling the construction of three-dimensional tissue structures, which could potentially be used to repair tissues in degenerative conditions such as osteoarthritis. The latter two applications would also benefit the public sector, in particular the Defence Science and Technology Laboratory, which supports research into both medical diagnostic devices and regenerative medicine. The UK aims to achieve a £10bn synbio market by 2030 (UK Synthetic Biology Strategic Plan 2016); the output from this proposal will simplify the implementation of synbio in eukaryotic cells and help to realise its promised economic benefits. During the Fellowship, I aim to establish at least one industrial partnership capable of commercialising applications of this research.

We anticipate that the general public will be the long-term beneficiary of this research, particularly in the area of healthcare. We will conduct public engagement activities throughout the Fellowship, to communicate outputs of our research with the general public and raise awareness of synbio. We will also monitor the public perception of specific applications of this research (e.g. using the tool to control human cell division), which will enable us to respond to shifts in public opinion and maximise its societal benefit. Relevant outcomes from this research will be used to shape the education and training strategies of professional bodies, such as the Biochemical Society, in relation to synbio. They will also be used to formulate responses to government consultations and enquiries on synbio, which will help to shape UK research, innovation and regulatory policies.