Isabelle AUJARD, Jean-Bernard BAUDIN, Arnaud GAUTIER, Ludovic JULLIEN, Thomas LE SAUX, Marie-Aude PLAMONT,
The hybridization of small synthetic molecular probes and genetically encoded modules opens exciting prospects to specifically target exogenous bioorthogonal functions in living systems. Genetic encoding ensures selective targeting in complex living systems, while the properties of exogenous molecules can be finely tailored by molecular engineering. We aim at developing hybrid reporters and biosensors for interrogating biological systems in all their complexity, focusing on the following challenges: (i) pushing multiplexed imaging to increase the number of targets one can image in a single experiment; (ii) pushing super-resolution microscopy to visualize biological processes in real-time at nanometric resolution; (iii) deciphering dynamic biological processes through the observation of protein-protein interactions, endogenous proteins, nucleic acids, enzymatic activities or cellular analytes. Our approach will consist in developing fluorogenic chromophores (so-called fluorogens) able to light up upon selective interaction with evolved genetically encoded modules acting as reporters or sensors. Molecular engineering will enable us, on one hand, to engineer the electronic structure of the chromophores to tune their spectral properties, or their physicochemical properties to modulate their cellular or physiological localization. Directed evolution approaches, on the other hand, will allow us to design specific fluorogen-activating proteic modules with unprecedented selectivity and performance through iterative rounds of genetic diversification and selection. The ability of conditioning fluorogen binding, and thus fluorescence, to a given recognition event or cellular signal will open great prospects for the development of probes and biosensors for basic and biomedical research.