SUBCELLULAR IMAGING OF DYNAMIC PROTEIN INTERACTIONS BY BIOLUMINESCENCE RESONANCE ENERGY TRANSFER

¹ IGF
² Universite de Montreal

^* To whom correspondence should be addressed. E-mail: julie.perroy{at}igf.cnrs.fr.

Submitted on July 12, 2007
Revised on August 16, 2007
Accepted on 24 September 2007

	Abstract

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 -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.

Key Words: BRET imaging, beta-arrestin recruitment, dynamic spatio-temporal protein-protein interactions, living cells, setting up experimental conditions

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