ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling

Abigail Harris; Pam Siggers; Silvia Corrochano; Nick Warr; Danielle Sagar; Daniel T. Grimes; Makoto Suzuki; Rebecca D. Burdine; Feng Cong; Bon Kyoung Koo; Hans Clevers; Isabelle Stévant; Serge Nef; Sara Wells; Raja Brauner; Bochra Ben Rhouma; Neïla Belguith; Caroline Eozenou; Joelle Bignon-Topalovic; Anu Bashamboo; Ken McElreavey; Andy Greenfield

doi:10.1073/pnas.1801223115

ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling

Abigail Harris, Pam Siggers, Silvia Corrochano, Nick Warr, Danielle Sagar, Daniel T. Grimes, Makoto Suzuki, Rebecca D. Burdine, Feng Cong, Bon Kyoung Koo, Hans Clevers, Isabelle Stévant, Serge Nef, Sara Wells, Raja Brauner, Bochra Ben Rhouma, Neïla Belguith, Caroline Eozenou, Joelle Bignon-Topalovic, Anu BashambooKen McElreavey, Andy Greenfield

Research output: Contribution to journal › Article › peer-review

65 Scopus citations

Abstract

Mammalian sex determination is controlled by the antagonistic interactions of two genetic pathways: The SRY-SOX9-FGF9 network promotes testis determination partly by opposing proovarian pathways, while RSPO1/WNT-β-catenin/FOXL2 signals control ovary development by inhibiting SRY-SOX9-FGF9. The molecular basis of this mutual antagonism is unclear. Here we show that ZNRF3, a WNT signaling antagonist and direct target of RSPO1-mediated inhibition, is required for sex determination in mice. XY mice lacking ZNRF3 exhibit complete or partial gonadal sex reversal, or related defects. These abnormalities are associated with ectopic WNT/ β-catenin activity and reduced Sox9 expression during fetal sex determination. Using exome sequencing of individuals with 46, XY disorders of sex development, we identified three human ZNRF3 variants in very rare cases of XY female presentation. We tested two missense variants and show that these disrupt ZNRF3 activity in both human cell lines and zebrafish embryo assays. Our data identify a testis-determining function for ZNRF3 and indicate a mechanism of direct molecular interaction between two mutually antagonistic organogenetic pathways.

Original language	English
Pages (from-to)	5474-5479
Number of pages	6
Journal	Proceedings of the National Academy of Sciences of the United States of America
Volume	115
Issue number	21
DOIs	https://doi.org/10.1073/pnas.1801223115
State	Published - 22 May 2018
Externally published	Yes

Bibliographical note

Publisher Copyright:
© 2018 National Academy of Sciences. All Rights Reserved.

Funding

We thank the husbandry team in Ward 5 of the Mary Lyon Centre at Harwell and the Frozen Embryo and Sperm Archive (FESA) and histology teams. We thank Dagmar Wilhelm for the kind gift of anti-FOXL2 antibody. We thank Phil Johnson for zebrafish husbandry. We acknowledge European Cooperation in Science & Technology (COST) Action BM1303 (DSDnet). This work was supported by the Medical Research Council by core funding Grant MC_U142684167 (to A.G.) at the Harwell Institute, and the Agence Nationale de la Recherche Grant ANR-10-LABX-73 (to K.M.). S.N. acknowledges support from Swiss National Science Foundation Grant 31003A_173070. M.S. was a visiting scientist supported by the Strategic International Research Exchange Program between Princeton University and National Institutes of Natural Sciences, Japan. D.T.G. was supported by National Institute of Arthritis and Mucoskeletal and Skin Diseases Pathway to Independence Award 1K99AR070905. Work in the R.D.B. laboratory is supported by the National Institute of Child Health and Development Grant 2R01HD048584. ACKNOWLEDGMENTS. We thank the husbandry team in Ward 5 of the Mary Lyon Centre at Harwell and the Frozen Embryo and Sperm Archive (FESA) and histology teams. We thank Dagmar Wilhelm for the kind gift of anti-FOXL2 antibody. We thank Phil Johnson for zebrafish husbandry. We acknowledge European Cooperation in Science & Technology (COST) Action BM1303 (DSDnet). This work was supported by the Medical Research Council by core funding Grant MC_U142684167 (to A.G.) at the Harwell Institute, and the Agence Nationale de la Recherche Grant ANR-10-LABX-73 (to K.M.). S.N. acknowledges support from Swiss National Science Foundation Grant 31003A_173070. M.S. was a visiting scientist supported by the Strategic International Research Exchange Program between Princeton University and National Institutes of Natural Sciences, Japan. D.T.G. was supported by National Institute of Arthritis and Mucoskeletal and Skin Diseases Pathway to Independence Award 1K99AR070905. Work in the R.D.B. laboratory is supported by the National Institute of Child Health and Development Grant 2R01HD048584.

Funders	Funder number
Harwell Institute
National Institute of Arthritis and Mucoskeletal and Skin Diseases Pathway
S.N.
National Institute of Arthritis and Musculoskeletal and Skin Diseases	K99AR070905
National Institute of Child Health and Human Development	2R01HD048584
Center for Outcomes Research and Evaluation, Yale School of Medicine
Princeton University
M.S.I. Foundation
Medical Research Council	MC_U142684167
European Cooperation in Science and Technology	COST, BM1303
Agence Nationale de la Recherche	ANR-10-LABX-73
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung	31003A_173070
National Institutes of Natural Sciences

Keywords

DSD
Organogenesis
Sex determination
WNT signaling
ZNRF3

Access to Document

10.1073/pnas.1801223115

Cite this

Harris, A., Siggers, P., Corrochano, S., Warr, N., Sagar, D., Grimes, D. T., Suzuki, M., Burdine, R. D., Cong, F., Koo, B. K., Clevers, H., Stévant, I., Nef, S., Wells, S., Brauner, R., Rhouma, B. B., Belguith, N., Eozenou, C., Bignon-Topalovic, J., ... Greenfield, A. (2018). ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling. Proceedings of the National Academy of Sciences of the United States of America, 115(21), 5474-5479. https://doi.org/10.1073/pnas.1801223115

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abstract = "Mammalian sex determination is controlled by the antagonistic interactions of two genetic pathways: The SRY-SOX9-FGF9 network promotes testis determination partly by opposing proovarian pathways, while RSPO1/WNT-β-catenin/FOXL2 signals control ovary development by inhibiting SRY-SOX9-FGF9. The molecular basis of this mutual antagonism is unclear. Here we show that ZNRF3, a WNT signaling antagonist and direct target of RSPO1-mediated inhibition, is required for sex determination in mice. XY mice lacking ZNRF3 exhibit complete or partial gonadal sex reversal, or related defects. These abnormalities are associated with ectopic WNT/ β-catenin activity and reduced Sox9 expression during fetal sex determination. Using exome sequencing of individuals with 46, XY disorders of sex development, we identified three human ZNRF3 variants in very rare cases of XY female presentation. We tested two missense variants and show that these disrupt ZNRF3 activity in both human cell lines and zebrafish embryo assays. Our data identify a testis-determining function for ZNRF3 and indicate a mechanism of direct molecular interaction between two mutually antagonistic organogenetic pathways.",

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Harris, A, Siggers, P, Corrochano, S, Warr, N, Sagar, D, Grimes, DT, Suzuki, M, Burdine, RD, Cong, F, Koo, BK, Clevers, H, Stévant, I, Nef, S, Wells, S, Brauner, R, Rhouma, BB, Belguith, N, Eozenou, C, Bignon-Topalovic, J, Bashamboo, A, McElreavey, K & Greenfield, A 2018, 'ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling', Proceedings of the National Academy of Sciences of the United States of America, vol. 115, no. 21, pp. 5474-5479. https://doi.org/10.1073/pnas.1801223115

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AU - Harris, Abigail

AU - Siggers, Pam

AU - Corrochano, Silvia

AU - Warr, Nick

AU - Sagar, Danielle

AU - Grimes, Daniel T.

AU - Suzuki, Makoto

AU - Burdine, Rebecca D.

AU - Cong, Feng

AU - Koo, Bon Kyoung

AU - Clevers, Hans

AU - Stévant, Isabelle

AU - Nef, Serge

AU - Wells, Sara

AU - Brauner, Raja

AU - Rhouma, Bochra Ben

AU - Belguith, Neïla

AU - Eozenou, Caroline

AU - Bignon-Topalovic, Joelle

AU - Bashamboo, Anu

AU - McElreavey, Ken

AU - Greenfield, Andy

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ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling

Abstract

Bibliographical note

Funding

Keywords

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