The Molecular Control of Calcitonin Receptor Signaling

Emma dal Maso, Alisa Glukhova, Yue Zhu, Javier Garcia-Nafria, Christopher G. Tate, Silvia Atanasio, Christopher A. Reynolds, Erney Ramirez-Aportela, Jose-Maria Carazo, Caroline A. Hick, Sebastian G. B. Furness, Debbie L. Hay, Yi-Lynn Liang, Laurence J. Miller, Arthur Christopoulos, Ming-Wei Wang, Denise Wootten, Patrick M. Sexton

Research output: Contribution to journalArticleResearchpeer-review

Abstract

The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 Å structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., et al. Phase-plate cryo- EM structure of a class B GPCR-G protein complex. Nature2017, 546, 118–123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 Å map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily.
Original languageEnglish
Pages (from-to)31-51
Number of pages21
JournalACS Pharmacology and Translational Sciences
Volume2
Issue number1
DOIs
Publication statusPublished - 11 Jan 2019

Cite this

dal Maso, Emma ; Glukhova, Alisa ; Zhu, Yue ; Garcia-Nafria, Javier ; Tate, Christopher G. ; Atanasio, Silvia ; Reynolds, Christopher A. ; Ramirez-Aportela, Erney ; Carazo, Jose-Maria ; Hick, Caroline A. ; Furness, Sebastian G. B. ; Hay, Debbie L. ; Liang, Yi-Lynn ; Miller, Laurence J. ; Christopoulos, Arthur ; Wang, Ming-Wei ; Wootten, Denise ; Sexton, Patrick M. / The Molecular Control of Calcitonin Receptor Signaling. In: ACS Pharmacology and Translational Sciences. 2019 ; Vol. 2, No. 1. pp. 31-51.
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title = "The Molecular Control of Calcitonin Receptor Signaling",
abstract = "The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 {\AA} structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., et al. Phase-plate cryo- EM structure of a class B GPCR-G protein complex. Nature2017, 546, 118–123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 {\AA} map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily.",
author = "{dal Maso}, Emma and Alisa Glukhova and Yue Zhu and Javier Garcia-Nafria and Tate, {Christopher G.} and Silvia Atanasio and Reynolds, {Christopher A.} and Erney Ramirez-Aportela and Jose-Maria Carazo and Hick, {Caroline A.} and Furness, {Sebastian G. B.} and Hay, {Debbie L.} and Yi-Lynn Liang and Miller, {Laurence J.} and Arthur Christopoulos and Ming-Wei Wang and Denise Wootten and Sexton, {Patrick M.}",
year = "2019",
month = "1",
day = "11",
doi = "10.1021/acsptsci.8b00056",
language = "English",
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pages = "31--51",
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dal Maso, E, Glukhova, A, Zhu, Y, Garcia-Nafria, J, Tate, CG, Atanasio, S, Reynolds, CA, Ramirez-Aportela, E, Carazo, J-M, Hick, CA, Furness, SGB, Hay, DL, Liang, Y-L, Miller, LJ, Christopoulos, A, Wang, M-W, Wootten, D & Sexton, PM 2019, 'The Molecular Control of Calcitonin Receptor Signaling', ACS Pharmacology and Translational Sciences, vol. 2, no. 1, pp. 31-51. https://doi.org/10.1021/acsptsci.8b00056

The Molecular Control of Calcitonin Receptor Signaling. / dal Maso, Emma; Glukhova, Alisa; Zhu, Yue; Garcia-Nafria, Javier; Tate, Christopher G.; Atanasio, Silvia; Reynolds, Christopher A.; Ramirez-Aportela, Erney; Carazo, Jose-Maria; Hick, Caroline A.; Furness, Sebastian G. B.; Hay, Debbie L.; Liang, Yi-Lynn; Miller, Laurence J.; Christopoulos, Arthur; Wang, Ming-Wei; Wootten, Denise; Sexton, Patrick M.

In: ACS Pharmacology and Translational Sciences, Vol. 2, No. 1, 11.01.2019, p. 31-51.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - The Molecular Control of Calcitonin Receptor Signaling

AU - dal Maso, Emma

AU - Glukhova, Alisa

AU - Zhu, Yue

AU - Garcia-Nafria, Javier

AU - Tate, Christopher G.

AU - Atanasio, Silvia

AU - Reynolds, Christopher A.

AU - Ramirez-Aportela, Erney

AU - Carazo, Jose-Maria

AU - Hick, Caroline A.

AU - Furness, Sebastian G. B.

AU - Hay, Debbie L.

AU - Liang, Yi-Lynn

AU - Miller, Laurence J.

AU - Christopoulos, Arthur

AU - Wang, Ming-Wei

AU - Wootten, Denise

AU - Sexton, Patrick M.

PY - 2019/1/11

Y1 - 2019/1/11

N2 - The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 Å structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., et al. Phase-plate cryo- EM structure of a class B GPCR-G protein complex. Nature2017, 546, 118–123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 Å map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily.

AB - The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 Å structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., et al. Phase-plate cryo- EM structure of a class B GPCR-G protein complex. Nature2017, 546, 118–123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 Å map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily.

U2 - 10.1021/acsptsci.8b00056

DO - 10.1021/acsptsci.8b00056

M3 - Article

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EP - 51

JO - ACS Pharmacology and Translational Sciences

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SN - 2575-9108

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