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Research Themes Drug discovery

GPCR subunits: Separate but not equal

PSI-SGKB [doi:10.1038/fa_psisgkb.2009.39]
Featured Article - September 2009
Short description: A functional complementation assay reveals that maximal heterotrimeric G-protein activation is achieved by agonist binding to one subunit of a dopamine D2 receptor dimer.

Cartoon of different D2R dimer activation states, with activation data for these states, from the perspective of agonist-mediated activation of protomer A.

What is the minimal functional unit of a G protein-coupled receptor (GPCR)? Some receptors have been proposed to exist in a 2:1 stoichiometry with heterotrimeric G proteins, although rhodopsin and the β2 adrenergic receptor can activate G proteins in vitro as monomers. In addition, it is not always clear whether agonists bind one or both subunits of a receptor dimer. Reporting in Nature Chemical Biology, Jonathan Javitch and colleagues use a functional complementation assay to study the stoichiometry of human dopamine D2 receptor (D2R) signaling. They find that a D2R dimer binds to a single heterotrimeric G protein and is maximally activated by the binding of an agonist to one receptor protomer.

To study D2R activation, Han et al. fused one D2R (called protomer B) to a novel pertussis-toxin-insensitive Gqi5 chimera that produced calcium-dependent luminescence upon activation. The fusion protein did not signal in response to agonist binding because the very short linker did not allow the G protein to couple to the receptor to which it was fused. However, co-expression of a D2R protomer not fused to a G protein (called protomer A) caused robust agonist-mediated activation. Therefore, agonist-induced D2R signaling is mediated by two receptor protomers and one G protein.

A mutation in protomer A that inhibited agonist binding blocked G-protein-mediated signaling. Surprisingly, this same mutation in protomer B increased G-protein activation compared to the wild-type homodimer. Inverse agonist binding to protomer B also enhanced activation. Thus, the binding of an agonist to one subunit of a dimer is necessary and sufficient for G-protein activation. Intriguingly, agonist binding to protomer B, as well as a constitutively active version of protomer B, both diminished agonist-induced G-protein activation, suggesting that the active conformation of protomer B inhibits signaling and that agonist binding per se is not required for this effect.

Computational modeling and additional mutagenesis studies suggested that the second intracellular loop of both protomers makes contact with the G protein. However, the third intracellular loop of protomer A but not protomer B is required for G-protein activation, which indicates that each protomer in a receptor dimer has a discrete function. Given the apparent asymmetrical role of these protomers and the importance of conformational changes in modulating G-protein activation, it will be interesting to determine the effects of heterodimerization on agonist-stimulated signaling. The complementation assay described in this report will be a useful technique for these future studies.

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  1. Y. Han et al. Allosteric communication between protomers of dopamine class A GPCR dimers modulates activation.
    Nature Chem. Biol. (2009). doi:10.1038/nchembio.199

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