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Inactive-state preassembly of Gq-coupled receptors and Gq heterotrimers

Abstract

G protein–coupled receptors (GPCRs) transmit signals by forming active-state complexes with heterotrimeric G proteins. It has been suggested that some GPCRs also assemble with G proteins before ligand-induced activation and that inactive-state preassembly facilitates rapid and specific G protein activation. However, no mechanism of preassembly has been described, and no functional consequences of preassembly have been demonstrated. Here we show that M3 muscarinic acetylcholine receptors (M3R) form inactive-state complexes with Gq heterotrimers in intact cells. The M3R C terminus is sufficient, and a six-amino-acid polybasic sequence distal to helix 8 (565KKKRRK570) is necessary for preassembly with Gq. Replacing this sequence with six alanine residues prevents preassembly, slows the rate of Gq activation and decreases steady-state agonist sensitivity. That other Gq-coupled receptors possess similar polybasic regions and also preassemble with Gq suggests that these GPCRs may use a common preassembly mechanism to facilitate activation of Gq heterotrimers.

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Figure 1: Immobile C-M3R decreases Gαq-V mobility.
Figure 2: Receptor activity does not affect Gαq-V mobility in intact cells.
Figure 3: The M3R C terminus is necessary and sufficient for preassembly with Gq.
Figure 4: A polybasic region in the M3R C terminus is necessary for preassembly with Gq.
Figure 5: Preassembly involves an electrostatic interaction and requires an electronegative plasma membrane.
Figure 6: A polybasic region in the M3R C terminus is necessary for efficient activation of Gq.

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Acknowledgements

We thank G. Digby and S. Kuravi for invaluable assistance. This work was supported by US National Institutes of Health grants GM076167 (G.W.), GM096762 and GM078319 (N.A.L.).

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K.Q. designed and performed experiments and analyzed data; C.D. performed experiments; G.W. designed experiments and analyzed data; N.A.L. designed experiments, analyzed data and wrote the paper.

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Correspondence to Nevin A Lambert.

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The authors declare no competing financial interests.

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Qin, K., Dong, C., Wu, G. et al. Inactive-state preassembly of Gq-coupled receptors and Gq heterotrimers. Nat Chem Biol 7, 740–747 (2011). https://doi.org/10.1038/nchembio.642

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