Monitoring lipid-protein interactions in planta using Förster resonance energy transfer.

Abstract

Phospholipids are not only the major structural components of cellular membranes but also important signaling molecules regulating various cellular and physiological processes. One mode of action by lipid mediators is via lipid-protein interactions to modulate the downstream cellular events. An increasing number of lipid-binding proteins have been identified using in vitro lipid-protein binding assays, but it has been challenging to monitor lipid-protein interactions in vivo. Here we describe one Förster resonance energy transfer (FRET)-based method using the cyan fluorescence protein (CFP)-tagged protein cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) and TopFluor TMR-labeled lipid phosphatidic acid (PA) to monitor the lipid-protein interaction in planta. This approach permits detection of the subcellular localization of lipid-protein interactions and dynamics of the interactions in planta in response to different cues.