Cryo-EM structure of constitutively active human Frizzled 7 in complex with heterotrimeric Gs

doi:10.1038/s41422-021-00525-6

. 2021 Dec;31(12):1311-1314.

doi: 10.1038/s41422-021-00525-6. Epub 2021 Jul 8.

Cryo-EM structure of constitutively active human Frizzled 7 in complex with heterotrimeric G_s

Lu Xu^#^{1

2

3

4}, Bo Chen^#¹, Hannes Schihada^#⁵, Shane C Wright^#^{5

6}, Ainoleena Turku^{5

7}, Yiran Wu¹, Gye-Won Han⁸, Maria Kowalski-Jahn⁵, Pawel Kozielewicz⁵, Carl-Fredrik Bowin⁵, Xianjun Zhang^{1

2

3

4

8}, Chao Li^{1

2

3

4}, Michel Bouvier⁶, Gunnar Schulte⁹, Fei Xu^{10

11

12

13}

Affiliations

¹ iHuman Institute, ShanghaiTech University, Shanghai, China.
² School of Life Science and Technology, ShanghaiTech University, Shanghai, China.
³ Center for Excellence in Molecular Cell Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
⁴ University of Chinese Academy of Sciences, Beijing, China.
⁵ Section of Receptor Biology & Signaling, Department of Physiology & Pharmacology, Karolinska Institutet, Stockholm, Sweden.
⁶ Institute for Research in Immunology and Cancer, Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, QC, Canada.
⁷ Orion Pharma R&D, Espoo, Finland.
⁸ Departments of Biological Sciences and Chemistry, Bridge Institute, University of Southern California, Los Angeles, CA, USA.
⁹ Section of Receptor Biology & Signaling, Department of Physiology & Pharmacology, Karolinska Institutet, Stockholm, Sweden. gunnar.schulte@ki.se.
¹⁰ iHuman Institute, ShanghaiTech University, Shanghai, China. xufei@shanghaitech.edu.cn.
¹¹ School of Life Science and Technology, ShanghaiTech University, Shanghai, China. xufei@shanghaitech.edu.cn.
¹² Center for Excellence in Molecular Cell Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China. xufei@shanghaitech.edu.cn.
¹³ University of Chinese Academy of Sciences, Beijing, China. xufei@shanghaitech.edu.cn.

^# Contributed equally.

PMID: 34239071
PMCID: PMC8648716
DOI: 10.1038/s41422-021-00525-6

Cryo-EM structure of constitutively active human Frizzled 7 in complex with heterotrimeric G_s

Lu Xu et al. Cell Res. 2021 Dec.

. 2021 Dec;31(12):1311-1314.

doi: 10.1038/s41422-021-00525-6. Epub 2021 Jul 8.

Authors

Affiliations

¹ iHuman Institute, ShanghaiTech University, Shanghai, China.
² School of Life Science and Technology, ShanghaiTech University, Shanghai, China.
³ Center for Excellence in Molecular Cell Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
⁴ University of Chinese Academy of Sciences, Beijing, China.
⁵ Section of Receptor Biology & Signaling, Department of Physiology & Pharmacology, Karolinska Institutet, Stockholm, Sweden.
⁶ Institute for Research in Immunology and Cancer, Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, QC, Canada.
⁷ Orion Pharma R&D, Espoo, Finland.
⁸ Departments of Biological Sciences and Chemistry, Bridge Institute, University of Southern California, Los Angeles, CA, USA.
⁹ Section of Receptor Biology & Signaling, Department of Physiology & Pharmacology, Karolinska Institutet, Stockholm, Sweden. gunnar.schulte@ki.se.
¹⁰ iHuman Institute, ShanghaiTech University, Shanghai, China. xufei@shanghaitech.edu.cn.
¹¹ School of Life Science and Technology, ShanghaiTech University, Shanghai, China. xufei@shanghaitech.edu.cn.
¹² Center for Excellence in Molecular Cell Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China. xufei@shanghaitech.edu.cn.
¹³ University of Chinese Academy of Sciences, Beijing, China. xufei@shanghaitech.edu.cn.

^# Contributed equally.

PMID: 34239071
PMCID: PMC8648716
DOI: 10.1038/s41422-021-00525-6

No abstract available

PubMed Disclaimer

Conflict of interest statement

M.B. is the president of the scientific advisory board for Domain Therapeutics. M.B. has filed patent applications related to some of the biosensors used in this work and the technology has been licensed to Domain Therapeutics.

Figures

**Fig. 1. Structure of constitutively active FZD₇ in complex with heterotrimeric mG_s.**
a Normalized BRET₀ values of ΔFZD_1–10 HEK293 cells transiently co-transfected with the G_s BRET sensor along with either negative control (mock), the β₂-adrenoceptor (β₂AR) or FZD₇. Data are represented as the means ± SEM of raw BRET₀ that were obtained from simple linear regression of five independent experiments measured in quadruplicates shown in Supplementary information, Fig. S1A and normalized to the negative control. *P < 0.05; **P < 0.01 (one-way ANOVA followed by Sidak’s multiple comparison). b An annotated 2D class average of FZD₇–mG_s–Nb35 complex. c Overall density map and atomic model of FZD₇–mG_s–Nb35 complex (CRD was omitted due to linker flexibility). FZD₇, blue; mGα_s, orange; Gβ, green; Gγ, yellow; Nb35, gray. d Insertion of the α5-helix (mGα_s, orange) into FZD₇ helical bundle represented as surface (ICL1, blue; ICL2, pink; ICL3, yellow; TM7/H8, green). e Schematics of interactions between FZD₇ and α5-helix. Hydrogen bonds are shown as red dashed lines. The red circle represents the hydrophobic interaction network. Yellow shades indicate residues that reside in TM5/6/ICL3; pink, TM3/4/ICL2; blue, TM1/2/ICL1; green, TM7/H8. f Superposition of FZD₇ (blue) and FZD₄ (yellow) structures, viewed from the intracellular side (bottom view). g Superposition of the active FZD₇ structure (blue) with the inactive FZD₄ (PDB: 6BD4, yellow), inactive FZD₅ (PDB: 6WW2, light pink), active SMO (PDB: 6OT0, gray) and inactive SMO (PDB: 5V57, green) structures. h Comparison of the cytoplasmic portion of TM6 (from K^6.28 to P^6.43) in FZD₇, FZD₄, active SMO and inactive SMO structures. i R^6.32, F^6.36, W^7.55 network in FZD₇, FZD₄ and active SMO structures. Blue dashed lines indicate the distance of F^6.36–W^7.55 and F^6.36–R^6.32 in FZD₇. Gray dashed lines indicate the distance of R^6.32–W^7.55 and W^7.55–F^6.36 in active SMO structure. j Normalized BRET₀ values of ΔFZD_1-10 HEK293 cells transiently co-transfected with rGFP-CAAX and Gα_s-67-RlucII, along with either negative control (mock), wild-type FZD_7, ΔCRD-FZD₇ or the indicated FZD₇ mutants. Data are represented as the means ± SEM of raw BRET₀ that were obtained from simple linear regression of four independent experiments measured in quadruplicates shown in Supplementary information, Fig. S11b and normalized to the negative control. **P < 0.01; ***P < 0.001 (one-way ANOVA followed by Tukey’s multiple comparison). k Normalized FRET₀ values of ΔFZD_1–10 HEK293 cells transiently co-transfected with the FRET-based cAMP biosensor along with either negative control (mock), wild-type FZD_7, ΔCRD-FZD₇ or the indicated FZD₇ mutants. Data are represented as the means ± SEM of raw FRET₀ that were obtained from simple linear regression of five independent experiments measured in quadruplicates shown in Supplementary information, Fig. S11c and normalized to the negative control. **P < 0.01; ****P < 0.0001 (one-way ANOVA followed by Sidak’s multiple comparison).

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References

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[1] Schulte G, Wright SC. Trends Pharmacol. Sci. 2018;39:828–42. doi: 10.1016/j.tips.2018.07.001. - DOI - PubMed

[2] Schulte G, Wright SC. Trends Pharmacol. Sci. 2018;39:828–42. doi: 10.1016/j.tips.2018.07.001. - DOI - PubMed

[3] Taipale J, Beachy PA. Nature. 2001;411:349–54. doi: 10.1038/35077219. - DOI - PubMed

[4] Taipale J, Beachy PA. Nature. 2001;411:349–54. doi: 10.1038/35077219. - DOI - PubMed

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[7] Phesse, T., Flanagan, D. & Vincan, E. Cancers8, 10.3390/cancers8050050 (2016). - PMC - PubMed

[8] Phesse, T., Flanagan, D. & Vincan, E. Cancers8, 10.3390/cancers8050050 (2016). - PMC - PubMed

[9] Nehmé R, Carpenter B, Singhal A, Strege A, Edwards PC, White CF, et al. PLoS ONE. 2017;12:e0175642. doi: 10.1371/journal.pone.0175642. - DOI - PMC - PubMed

[10] Nehmé R, Carpenter B, Singhal A, Strege A, Edwards PC, White CF, et al. PLoS ONE. 2017;12:e0175642. doi: 10.1371/journal.pone.0175642. - DOI - PMC - PubMed

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Cryo-EM structure of constitutively active human Frizzled 7 in complex with heterotrimeric G_s

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Cryo-EM structure of constitutively active human Frizzled 7 in complex with heterotrimeric G_s

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