Delivering the impossible - novel fatty acid delivery reagents to enable in vivo research and discovery

Fatty acids are essential parts of all living things and form the main part of fats and oils. They are also key components of the function of every cell. Every cell uses fatty acids in their membrane lipids and also uses them to alter the function or role of proteins within the cell. We are particularly interested in understanding how fatty acids are used in this way in plants in a process called S-acylation.

Recently, some chemical tools that look like fatty acids but have a reactive alkyne "handle" attached, were shown to behave like normal fatty acids when given to some cell types. The alkyne handle allows researchers to isolate proteins which are attached to that fatty acid and perform specific analysis on them. Unfortunately, this method does not work outside of a limited number of specific cell types. This is probably because most cells aren't able to take up fatty acids that are supplied to them. This is particularly true in plants where there is no need to absorb fatty acids from the outside environment as they can make their own. To make matters worse, the chemical nature of fatty acids means that they are also not soluble in water which reduces the dose that can be given in each experiment. This in turn reduces the sensitivity of the experiment to detect proteins with fatty acids attached.

In this project we aim to make fatty acids with alkyne handles that are more soluble enabling them to get inside the cell without any effort on the part of the organism. This uses validated ideas from the 1980's coupled to cutting edge chemical technology. Using these chemical tools, we will be able to investigate a wide range of processes involving fatty acids in different organisms.

We will also share our tools with other researchers to enable them to do previously impossible experiments on fatty acid metabolism in their organisms of interest.

Fatty acid metabolism is an essential process common to all life, with fatty acids being both core components of cellular membranes and key regulators of protein function through fatty acid-based post-translational modifications such as S-acylation. Understanding fatty acid metabolism is also fundamental to many areas of biotechnological, health or industrial interest such as biofuels, plant seed oils, dietary fat composition, type-I diabetes, prostaglandin production or lipid storage diseases. The direct study of S-acylation in eukaryotes, outside of a very limited number of cell cultures, is blocked by the lack of effective research tools to incorporate, monitor and trace fatty acids. The broader fatty acid metabolism communities also urgently need resources to enable timely research progression and new ways of experimentation. With a focus on S-acylation in plants, this interdisciplinary work will combine existing validated knowledge to produce effective reagents for in vivo delivery of click-chemistry enabled fatty acid alkyne probes. Using Arabidopsis, we will validate these new reagents by profiling the dynamics of protein S-acylation and examining the plant secreted acylated proteome, a hitherto inaccessible area of cell-to-cell signalling in plants. In addition, through committed international collaborators, we will deploy these novel reagents to other recalcitrant systems such as mammalian primary cell cultures, intact organs and tissues. Following validation, we will make reagents freely available to the research community to assess utility across the wider fatty acid metabolism field. These universally applicable tools will service unmet global needs and enable fatty acid research across a far wider range of organisms and systems than is currently possible, delivering international impact, exceptional value for money and support BBSRC "technology development for the biosciences" priority.