BBSRC EASTBIO DTP - Novel chemical tools for non-invasive imaging of plant metabolism

Optical imaging and light microscopy have revolutionized the way biologists interrogate complex cellular systems with high resolution. Our group, as many others, has contributed to the development of fluorescence microscopy with the preparation of optical probes to visualize molecules inside cells with excellent specificity and sensitivity (Vendrell and Oparka et al. Plant Physiol., 2015). Although fluorescence microscopy is still the method of choice for cellular imaging, it is limited when it comes to metabolic studies because fluorescent labels are bulky structures and introduce considerable perturbation to small molecule metabolites.

Stimulated Raman Scattering (SRS) microscopy has emerged as a revolutionary imaging modality to overcome these limitations. Unlike fluorescence imaging, SRS requires tiny and Raman-active chemical groups (e.g., alkyne groups, stable isotopes, fluorine atoms) that can be readily introduced into small molecule metabolites to track them in live systems and in real time (Hulme et al. Chem. Soc. Rev. 2016).

In this project, our aim is to develop small Raman-active sucrose probes (i.e. sucrose analogues with simple single atom modifications, such as F or D atoms replacing O or H) and to optimise them to study sucrose metabolism in plants non-invasively and in real time. We will generate a chemical toolbox that will enable to image for the first time the transport and fate of sucrose in live plants using SRS microscopy. Furthermore, this project will create a myriad of opportunities in biological research by developing a revolutionary chemical biology platform to study the localisation and function of small molecule metabolites that have not been trackable to date with high spatial and temporal resolution.

Current tools to study the localisation and partitioning of sucrose in plants are limited to 14C-labelled probes, which are very limited in spatial resolution and not amenable to in vivo imaging. Our Raman-active sucrose probes will allow to study for the first time key processes associated with the transport and unloading of sucrose inside plants with subcellular resolution. This will open new opportunities in Agriculture and Food Security as we will be able to test the ability of sucrose transporters proteins to transport glucoside-based synthetic molecules, such as herbicides, into the phloem of plants (Vendrell and Oparka et al. Plant Physiol. 2015). We will also be able to validate sucrose transporters as means for targeting xenobiotics into pathogens, as these receptors regulate the uptake of plant’s sucrose and might constitute an ‘Achilles Heel’ in the life cycle of pathogens. With these new probes, we will be able to study the extent to which our probes cross host and pathogen membranes.

Vendrell and Oparka have been actively collaborating for the last 2 years in the development of chemical imaging tools to interrogate plant biology, and they are co-PIs in a BBSRC Industrial Partnership Award (BB/M025160/1) with the multinational Syngenta (2015-2018). Vendrell is an expert in chemical biology and Raman imaging (Vendrell et al. Trends Biotechnol. 2013), whereas Oparka is a world-leader in high-resolution imaging of phloem transport.

The student will be part of the highly multidisciplinary team led by Vendrell and Oparka, and he/she will receive training in chemistry (i.e. development and characterisation of suitable chemical probes), biology (i.e. optimisation of models and assays to study metabolism and transport in plants) and imaging (i.e. SRS, Raman and fluorescence microscopy). The student will also benefit from a strong collaborative network with complementary expertise: 1) through collaboration with Prof. John Ward (Minnesota), we will confirm that Raman-active sucrose probes are transported on sucrose carriers using yeast and oocyte analyses. Prof. Ward is an expert in this area and his lab is uniquely equipped for these analyses; 2) collaboration with the group of Dr Alan Serrels (Edinburgh) as an expert in high resolution imaging and SRS microscopy; 3) through collaboration with Dr. Tim Hawkes and Dr. Torquil Fraser at Syngenta (Jealott’s Hill, Bracknell), the student will be exposed to the diverse expertise of Syngenta in chemistry, xenobiotic movement and chemical screening, which will be essential to the success of this project, as well as the potential commercial application of such chemistry to field conditions.