Chemical genetics of the retrograde transport route – RETROscreen

The goal of this project is the identification and characterization of novel regulators of the retrograde transport route in mammalian cells. Many cellular functions depend on this trafficking pathway, such as the establishment of wnt morphogen gradients, the intracellular compartmentalization of signaling molecules and of transporters for glucose, copper and iron. The retrograde route has also attracted attention in the therapeutic field. It is used by numerous pathogens or pathogenic products to enter into cells, and retrograde transport appears to be misregulated in Alzheimer's disease. Biotechnological developments are underway for its exploitation for the intracellular delivery of therapeutic compounds such as antigenic proteins, nucleic acids, contrast agents, nanoparticles… By high throughput screening, we recently identified two compounds acting as inhibitors of the retrograde route (Stechmann et al., Cell 2010). These compounds selectively block the intracellular transport of toxins from plants and pathogenic bacteria, and one of these compounds potently protects mice against lethal challenges by the plant toxin ricin, a potential bioterror weapon. In the context of the current proposal, these small molecules or their photoactivable analogues will be labeled with tritium, or alternatively will be tagged with biotin. These tools will be used for the identification of the cellular targets of the inhibitors by chromatographic methods and mass spectrometry. The cellular functions and subcellular localization of these targets will be studied. We will particularly analyze how they regulate retrograde transport, and vice versa, whether and how retrograde transport is critical for their functions.