Abstract

It is now clear that multiple routes exist in cells to transport proteins and lipids toward various compartments. To analyse these routes, there is a strong a need for a versatile, highly synchronized assay, usable at physiological temperature in living cells. We developed the RUSH system (Retention Using Selective Hooks), a two-state assay based on the reversible interaction of a Reporter protein with a Hook protein stably localized in a donor compartment. This system can be used to study and quantify the trafficking of very diverse proteins of different size, shape and destined to various compartments. The RUSH system can be used for real time monitoring of transport in living cells as well as for for High Throughput screening using automated imaging.

We are using the RUSH assay at multiple scales to monitor the transport of proteins in cells and analyse their intracellular signatures, to map the diversity of trafficking routes in terms of biochemical composition, to integrate these routes with other cellular machineries, like cellular adhesion and polarization, and to screen diverse chemical libraries and identify potential therapeutic molecules.