A library of fluorescent probes to study Reactive Oxygen Species At the Single cell level – ROSAS

Billions of years of photosynthesis have resulted in an oxygen-rich atmosphere in which the respiratory chain and aerobic life could evolve. This allowed the apparition of so-called “superior life forms”. However, the use of oxygen as an electron acceptor results in the concomitant formation of partially reduced oxygen species, which are highly reactive, such as hydrogen peroxide, the superoxide anion, and the hydroxyl radical. These species, called Reactive Oxygen Species (ROS), induce oxidative lesions on biological macromolecules, eventually resulting (i) in death or (ii) in apparent loss in the ability to grow on plate for bacteria. Paradoxically, ROS, especially H2O2, have also signaling properties. To decipher ROS biological and toxicological impact requires being able to appreciate their abundance within the cell and its compartments.
This fundamental project will target the development of a new integrated collection of molecular probes (toolbox) for the study by in vivo imaging (epifluorescence microscopy) of the presence of ROS at a single cell level. This will involve design, synthesis, characterization and validation of new ROS-sensitive probes.
In terms of biological goal, we aim at a better understanding of how each bacteria cope with oxidative stress generated during the plating procedure, and how this response affects the cells fate during recovery on plate. We will use Escherichia coli/Legionella pneumophila as two model organisms for studying the basic mechanisms underlying the apparent loss of ability of viable bacteria to grow on plate.
Moreover, we believe that the tools and strategies that we will develop can also find applications in other fields of biological research, where specific ROS fluorescent probes could be useful. Indeed, several biological groups (in France and abroad) have already contacted Partner 2, in order to use future ROS fluorescent probes. After adequate protection, the new molecular tools will be made available to the scientific community by publication and, if relevant, commercialization. As observed via PubMed, more than 2000 publications are referenced (keywords : (hydrogen peroxide or superoxide) and fluorescent probe), with a clear exponential growth of the number of publications in the last 5 years, showing a clear interest in this field. Moreover, several companies (Molecular probes, Sigma Aldrich, Invitrogen, Cayman…) commercialize fluorescent probes against ROS, indicating also a clear economical interest. For all these reasons we believe that our project is both timely and of major interest.