SUBCELLULAR IMAGING OF DYNAMIC PROTEIN INTERACTIONS BY BIOLUMINESCENCE RESONANCE ENERGY TRANSFER.

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Coulon, Vincent | Audet, Martin | Homburger, Vincent | Bockaert, Joël | Fagni, Laurent | Bouvier, Michel | Perroy, Julie

Edité par CCSD ; Biophysical Society -

Despite the fact that numerous studies suggest the existence of receptor multiprotein complexes, visualization and monitoring of the dynamics of such protein assemblies remains a challenge. In the present study we established appropriate conditions to study spatio-temporally-resolved images of such protein assemblies using bioluminescence resonance energy transfer (BRET) in mammalian living cells. Using covalently linked renilla-luciferase and yellow fluorescent proteins, we depicted the time course of dynamic changes in the interaction between the V2-vasopressin receptor and beta-arrestin induced by a receptor agonist. The protein-protein interactions were resolved at the level of sub-cellular compartments (nucleus, plasma membrane or endocytic vesicules) and in real time within tens of seconds to tens of minutes time frame. These studies provide a proof of principle, as well as experimental parameters and controls required for high resolution dynamic studies using BRET imaging, in single cells.

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