TY - JOUR
T1 - Variable G protein determinants of GPCR coupling selectivity
AU - Okashah, Najeah
AU - Wan, Qingwen
AU - Ghosh, Soumadwip
AU - Sandhu, Manbir
AU - Inoue, Asuka
AU - Vaidehi, Nagarajan
AU - Lambert, Nevin A.
N1 - Funding Information:
ACKNOWLEDGMENTS. We thank Steve Ikeda, Jonathan Javitch, Kevin Pfleger, Philip Wedegaertner, Carl Johnson, and Bryan Roth for providing plasmid DNA used in this study. This work was supported by National Institutes of Health Grants GM130142 (to N.A.L.) and GM117923 (to N.V.) and Ruth L. Kirschstein National Research Service Award Individual Fellowship GM131672 (to N.O.). A.I. was funded by the Advanced Research & Development programs PRIME JP17gm5910013 and LEAP JP17gm0010004 from Japan Agency for Medical Research and Development, and a grant-in-aid for scientific research KAKENHI 17K08264 from the Japan Society for the Promotion of Science.
Publisher Copyright:
© 2019 National Academy of Sciences. All rights reserved.
PY - 2019/6/11
Y1 - 2019/6/11
N2 - G protein-coupled receptors (GPCRs) activate four families of heterotrimeric G proteins, and individual receptors must select a subset of G proteins to produce appropriate cellular responses. Although the precise mechanisms of coupling selectivity are uncertain, the Gα subunit C terminus is widely believed to be the primary determinant recognized by cognate receptors. Here, we directly assess coupling between 14 representative GPCRs and 16 Gα subunits, including one wild-type Gα subunit from each of the four families and 12 chimeras with exchanged C termini. We use a sensitive bioluminescence resonance energy transfer (BRET) assay that provides control over both ligand and nucleotide binding, and allows direct comparison across G protein families. We find that the Gs- and Gq-coupled receptors we studied are relatively promiscuous and always couple to some extent to Gi1 heterotrimers. In contrast, Gi-coupled receptors are more selective. Our results with Gα subunit chimeras show that the Gα C terminus is important for coupling selectivity, but no more so than the Gα subunit core. The relative importance of the Gα subunit core and C terminus is highly variable and, for some receptors, the Gα core is more important for selective coupling than the C terminus. Our results suggest general rules for GPCR-G protein coupling and demonstrate that the critical G protein determinants of selectivity vary widely, even for different receptors that couple to the same G protein.
AB - G protein-coupled receptors (GPCRs) activate four families of heterotrimeric G proteins, and individual receptors must select a subset of G proteins to produce appropriate cellular responses. Although the precise mechanisms of coupling selectivity are uncertain, the Gα subunit C terminus is widely believed to be the primary determinant recognized by cognate receptors. Here, we directly assess coupling between 14 representative GPCRs and 16 Gα subunits, including one wild-type Gα subunit from each of the four families and 12 chimeras with exchanged C termini. We use a sensitive bioluminescence resonance energy transfer (BRET) assay that provides control over both ligand and nucleotide binding, and allows direct comparison across G protein families. We find that the Gs- and Gq-coupled receptors we studied are relatively promiscuous and always couple to some extent to Gi1 heterotrimers. In contrast, Gi-coupled receptors are more selective. Our results with Gα subunit chimeras show that the Gα C terminus is important for coupling selectivity, but no more so than the Gα subunit core. The relative importance of the Gα subunit core and C terminus is highly variable and, for some receptors, the Gα core is more important for selective coupling than the C terminus. Our results suggest general rules for GPCR-G protein coupling and demonstrate that the critical G protein determinants of selectivity vary widely, even for different receptors that couple to the same G protein.
KW - G protein selectivity
KW - G protein-coupled receptor
KW - GPCR
KW - Ternary complex
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U2 - 10.1073/pnas.1905993116
DO - 10.1073/pnas.1905993116
M3 - Article
C2 - 31142646
AN - SCOPUS:85067191355
SN - 0027-8424
VL - 116
SP - 12054
EP - 12059
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 24
ER -