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First published online August 9, 2007
doi: 10.1242/10.1242/jcs.001404

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A FRET map of membrane anchors suggests distinct microdomains of heterotrimeric G proteins

Ecole Polytechnique Fédérale de Lausanne (EPFL), Institut des Sciences et Ingénierie Chimiques, CH-1015 Lausanne, Switzerland

e-mails: d.abankwa{at}imb.uq.edu.au; horst.vogel{at}epfl.ch

Accepted 21 June 2007

The standard model of heterotrimeric G protein signaling postulates a dissociation of G and G subunits after activation. We hypothesized that the different combination of lipid-modifications on G and G subunits directs them into different microdomains. By characterizing rapidly and at high sensitivity 38 fluorescence resonance energy transfer (FRET) pairs of heterotrimeric-G-protein constructs, we defined their microdomains in relation to each other, free from the constraints of the raft/non-raft dualism. We estimated that in a cell 30% of these membrane-anchored proteins are mostly clustered in 3400-16,200 copies of 30-nm microdomains. We found that the membrane anchors of G and G subunits of both the G_i/o and G_q family co-cluster differently with microdomain markers. Moreover, anchors of the G_i/o and G_q subunits co-clustered only weakly, whereas constructs that contained the anchors of the corresponding heterotrimers co-clustered considerably, suggesting the existence of at least three types of microdomain. Finally, FRET experiments with full-length heterotrimeric G proteins confirmed that the inactive, heterotrimerized G subunit is in microdomains shared by heterotrimers from different subclasses, from where it displaces upon activation into a membrane-anchor- and subclass-specific microdomain.

Key words: FRET, Heterotrimeric G protein, Microdomain, Nanodomain, Raft

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