Congratulations with the PhD Victor
On 9 October 2025, Victor de Prado Parralejo was awarded his PhD degree after defending his thesis titled "Untargeted assessment of membrane transporter substrate spectra" at the Faculty of Science. The defense took place in the presence of family, friends and colleagues, marking the culmination of his research journey.
Summary of the thesis
Transporters are protein shuttles that enable the movement of chemical compounds (metabolites) across biological membranes. Determining transporter substrate spectra (the range of metabolites it transports) is a fundamental step in determining its physiological function(s) and how this metabolic flow contributes to the chemical constitution of organisms. Finding the transporter(s) for predetermined metabolites of interest has been the predominant approach to exploring transport. Nevertheless, these targeted approaches have restricted our transport insight to a limited set of metabolites while leaving many transporters lacking identified substrates.
The overall aims of this thesis are to develop untargeted tools and strategies to expand and reassess the plant transporter substrate spectra. First, we develop an untargeted functional screen with a set of fourteen transporters by expressing them in unfertilized frog egg (oocyte) membranes and analyzing the translocation of metabolites from plant metabolic mixtures. We
reveal different substrate accumulation patterns according to the origin of the plant extracts, and we show the need to dilute these extracts so as not to compromise the membrane selectivity that transporter function provides. We both reflect the published substrate spectra for some of these transporters and suggest new substrates. Additionally, we report on substrate promiscuity for some transporters that nuances their links to development through phytohormone transport.
Next, we optimize the generation of screening mixtures. We explore the compatibility of extracts with oocytes according to the plant extraction procedure. We balance the capacity of different extraction protocols to capture the metabolic diversity of plants with their undesirable membrane-permeating properties. Furthermore, we increase the metabolic diversity of extracts by using plants treated with different stressors. We profile the metabolic diversity of a liquid extraction procedure that offers reproducibility, diversity, and oocyte compatibility.
Finally, we combine the data analysis insight generated from the initial screening with the optimized extract and put it to the test with 217 transporters from the model plant Arabidopsis thaliana. These transporters represent all the members of Arabidopsis thaliana's Major Facilitator Superfamily (MFS), the biggest superfamily of transporters in the plant kingdom. MFS transporters transport a variety of compounds from primary to specialized metabolites, and a sizeable portion of them have hardly been characterized. We scale the analysis pipeline to adapt to this first-of-a-kind effort in plant biology by developing tools and strategies to tackle and interpret this massive dataset. This thesis contains the first approach to the dataset by displaying the transporters that accumulate the highest number of substrates and offers the first evidence for the deorphanization of transporters that were never characterized.
This work lays the cornerstone for the untargeted screening of plant transportomes. The strategies developed here offer an approach for these massive datasets applicable to other organisms, as well as novel substrate-transporter links.
Principal supervisors:
Associate professor Hussam Nour-Eldin, Section for Molecular Plant Biology
Professor Meike Burow, Section for Plant Glyco Biologi
The assessment committee consisted of:
Assessment committee
Chairperson: Professor Rosa Laura Lopez Marques, Department of Plant and Environmental Sciences
Professor Stefan Broer, Membrane transport and nutrition, Australian National University, Canberra
Research Director, Benoit Lacombe Institute for Plant Sciences of Montpellier, France
Victor will continue as postdoc for another year at the DynaMo Center.