Single-cell, whole-embryo phenotyping of mammalian developmental disorders

Xingfan Huang; Jana Henck; Chengxiang Qiu; Varun K.A. Sreenivasan; Saranya Balachandran; Oana V. Amarie; Martin Hrabě de Angelis; Rose Yinghan Behncke; Wing Lee Chan; Alexandra Despang; Diane E. Dickel; Madeleine Duran; Annette Feuchtinger; Helmut Fuchs; Valerie Gailus-Durner; Natja Haag; Rene Hägerling; Nils Hansmeier; Friederike Hennig; Cooper Marshall; Sudha Rajderkar; Alessa Ringel; Michael Robson; Lauren M. Saunders; Patricia da Silva-Buttkus; Nadine Spielmann; Sanjay R. Srivatsan; Sascha Ulferts; Lars Wittler; Yiwen Zhu; Vera M. Kalscheuer; Daniel M. Ibrahim; Ingo Kurth; Uwe Kornak; Axel Visel; Len A. Pennacchio; David R. Beier; Cole Trapnell; Junyue Cao; Jay Shendure; Malte Spielmann

doi:10.1038/s41586-023-06548-w

Single-cell, whole-embryo phenotyping of mammalian developmental disorders

Xingfan Huang, Jana Henck, Chengxiang Qiu, Varun K.A. Sreenivasan, Saranya Balachandran, Oana V. Amarie, Martin Hrabě de Angelis, Rose Yinghan Behncke, Wing Lee Chan, Alexandra Despang, Diane E. Dickel, Madeleine Duran, Annette Feuchtinger, Helmut Fuchs, Valerie Gailus-Durner, Natja Haag, Rene Hägerling, Nils Hansmeier, Friederike Hennig, Cooper MarshallSudha Rajderkar, Alessa Ringel, Michael Robson, Lauren M. Saunders, Patricia da Silva-Buttkus, Nadine Spielmann, Sanjay R. Srivatsan, Sascha Ulferts, Lars Wittler, Yiwen Zhu, Vera M. Kalscheuer, Daniel M. Ibrahim, Ingo Kurth, Uwe Kornak, Axel Visel, Len A. Pennacchio, David R. Beier, Cole Trapnell, Junyue Cao^*, Jay Shendure^*, Malte Spielmann^*

^*Corresponding author for this work

Institute of Human Genetics

21 Citations (Scopus)

Abstract

Mouse models are a critical tool for studying human diseases, particularly developmental disorders¹. However, conventional approaches for phenotyping may fail to detect subtle defects throughout the developing mouse². Here we set out to establish single-cell RNA sequencing of the whole embryo as a scalable platform for the systematic phenotyping of mouse genetic models. We applied combinatorial indexing-based single-cell RNA sequencing³ to profile 101 embryos of 22 mutant and 4 wild-type genotypes at embryonic day 13.5, altogether profiling more than 1.6 million nuclei. The 22 mutants represent a range of anticipated phenotypic severities, from established multisystem disorders to deletions of individual regulatory regions^4,5. We developed and applied several analytical frameworks for detecting differences in composition and/or gene expression across 52 cell types or trajectories. Some mutants exhibit changes in dozens of trajectories whereas others exhibit changes in only a few cell types. We also identify differences between widely used wild-type strains, compare phenotyping of gain- versus loss-of-function mutants and characterize deletions of topological associating domain boundaries. Notably, some changes are shared among mutants, suggesting that developmental pleiotropy might be ‘decomposable’ through further scaling of this approach. Overall, our findings show how single-cell profiling of whole embryos can enable the systematic molecular and cellular phenotypic characterization of mouse mutants with unprecedented breadth and resolution.

Original language	English
Journal	Nature
Volume	623
Issue number	7988
Pages (from-to)	772-781
Number of pages	10
ISSN	0028-0836
DOIs	https://doi.org/10.1038/s41586-023-06548-w
Publication status	Published - 23.11.2023

Funding

We thank S. Mundlos and C. Prada for helpful discussions around data processing and analysis and interpretation of results; all members of the laboratories of J.C., J.S. and M.S. for continuous support and input; V. Suckow for genotyping the Ror2 -knock-in and Cdkl5 −/Y mice; and S. Houghtaling and T.-H. Ho for breeding and embryo collection of Ttc21b, Carm1 and Gli2 mice. N.Haag and I.K. thank M. Ebbinghaus for help with breeding of Scn11a -GOF mice. J.S. and work in the laboratory of J.S. were supported by the Paul G. Allen Frontiers Foundation (Allen Discovery Center grant to J.S. and C.T.), the National Institutes of Health (NIH; grant UM1HG011531 to J.S.), Alex’s Lemonade Stand’s Crazy 8 Initiative (to J.S.) and the Bonita and David Brewer Fellowship (C.Q.). Work at the E.O. Lawrence Berkeley National Laboratory was supported by US NIH grants to L.A.P. and A.V. (UM1HG009421 and R01HG003988) and carried out under US Department of Energy Contract DE-AC02-05CH11231, University of California. U.K. was supported by the Deutsche Forschungsgemeinschaft (DFG) (KO 2891/6-1) and ERA-Net for Research on Rare Diseases (EUROGLYCAN-omics). D.M.I. was supported by the Deutsche Forschungsgemeinschaft (DFG) (IB 139/1-1 and IB 139/6-1). I.K. was supported by the Deutsche Forschungsgemeinschaft (DFG) (KU1587/3-1, KU1587/10-1). R.H. was supported by the European Union (ERC, PREVENT, 101078827). J.S. is an Investigator of the Howard Hughes Medical Institute. M.S. is a DZHK principal investigator and is supported by grants from the Deutsche Forschungsgemeinschaft (DFG; SP1532/3-1, SP1532/4-1 and SP1532/5-1) and the Deutsches Zentrum für Luft- und Raumfahrt (DLR 01GM1925). D.R.B. was supported by R01HD36404 from the National Institute of Child Health and Human Development. J.C. is supported by the NIH (grant DP2 HG012522-01 and RM1HG011014) and the Rockefeller University. M.H.d.A. and work at the German Mouse Clinic were supported by the German Federal Ministry of Education and Research (Infrafrontier grant 01KX1012); German Center for Diabetes Research (DZD). We thank S. Mundlos and C. Prada for helpful discussions around data processing and analysis and interpretation of results; all members of the laboratories of J.C., J.S. and M.S. for continuous support and input; V. Suckow for genotyping the Ror2-knock-in and Cdkl5 −/Y mice; and S. Houghtaling and T.-H. Ho for breeding and embryo collection of Ttc21b, Carm1 and Gli2 mice. N.Haag and I.K. thank M. Ebbinghaus for help with breeding of Scn11a-GOF mice. J.S. and work in the laboratory of J.S. were supported by the Paul G. Allen Frontiers Foundation (Allen Discovery Center grant to J.S. and C.T.), the National Institutes of Health (NIH; grant UM1HG011531 to J.S.), Alex’s Lemonade Stand’s Crazy 8 Initiative (to J.S.) and the Bonita and David Brewer Fellowship (C.Q.). Work at the E.O. Lawrence Berkeley National Laboratory was supported by US NIH grants to L.A.P. and A.V. (UM1HG009421 and R01HG003988) and carried out under US Department of Energy Contract DE-AC02-05CH11231, University of California. U.K. was supported by the Deutsche Forschungsgemeinschaft (DFG) (KO 2891/6-1) and ERA-Net for Research on Rare Diseases (EUROGLYCAN-omics). D.M.I. was supported by the Deutsche Forschungsgemeinschaft (DFG) (IB 139/1-1 and IB 139/6-1). I.K. was supported by the Deutsche Forschungsgemeinschaft (DFG) (KU1587/3-1, KU1587/10-1). R.H. was supported by the European Union (ERC, PREVENT, 101078827). J.S. is an Investigator of the Howard Hughes Medical Institute. M.S. is a DZHK principal investigator and is supported by grants from the Deutsche Forschungsgemeinschaft (DFG; SP1532/3-1, SP1532/4-1 and SP1532/5-1) and the Deutsches Zentrum für Luft- und Raumfahrt (DLR 01GM1925). D.R.B. was supported by R01HD36404 from the National Institute of Child Health and Human Development. J.C. is supported by the NIH (grant DP2 HG012522-01 and RM1HG011014) and the Rockefeller University. M.H.d.A. and work at the German Mouse Clinic were supported by the German Federal Ministry of Education and Research (Infrafrontier grant 01KX1012); German Center for Diabetes Research (DZD).

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

SDG 3 Good Health and Well-being

Research Areas and Centers

Research Area: Medical Genetics

DFG Research Classification Scheme

2.22-03 Human Genetics
2.11-05 General Genetics and Functional Genome Biology

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10.1038/s41586-023-06548-w

Cite this

Huang, X., Henck, J., Qiu, C., Sreenivasan, V. K. A., Balachandran, S., Amarie, O. V., Hrabě de Angelis, M., Behncke, R. Y., Chan, W. L., Despang, A., Dickel, D. E., Duran, M., Feuchtinger, A., Fuchs, H., Gailus-Durner, V., Haag, N., Hägerling, R., Hansmeier, N., Hennig, F., ... Spielmann, M. (2023). Single-cell, whole-embryo phenotyping of mammalian developmental disorders. Nature, 623(7988), 772-781. https://doi.org/10.1038/s41586-023-06548-w

@article{db2d77a61fbe48b4b81e03544b4c3020,

title = "Single-cell, whole-embryo phenotyping of mammalian developmental disorders",

abstract = "Mouse models are a critical tool for studying human diseases, particularly developmental disorders1. However, conventional approaches for phenotyping may fail to detect subtle defects throughout the developing mouse2. Here we set out to establish single-cell RNA sequencing of the whole embryo as a scalable platform for the systematic phenotyping of mouse genetic models. We applied combinatorial indexing-based single-cell RNA sequencing3 to profile 101 embryos of 22 mutant and 4 wild-type genotypes at embryonic day 13.5, altogether profiling more than 1.6 million nuclei. The 22 mutants represent a range of anticipated phenotypic severities, from established multisystem disorders to deletions of individual regulatory regions4,5. We developed and applied several analytical frameworks for detecting differences in composition and/or gene expression across 52 cell types or trajectories. Some mutants exhibit changes in dozens of trajectories whereas others exhibit changes in only a few cell types. We also identify differences between widely used wild-type strains, compare phenotyping of gain- versus loss-of-function mutants and characterize deletions of topological associating domain boundaries. Notably, some changes are shared among mutants, suggesting that developmental pleiotropy might be {\textquoteleft}decomposable{\textquoteright} through further scaling of this approach. Overall, our findings show how single-cell profiling of whole embryos can enable the systematic molecular and cellular phenotypic characterization of mouse mutants with unprecedented breadth and resolution.",

author = "Xingfan Huang and Jana Henck and Chengxiang Qiu and Sreenivasan, \{Varun K.A.\} and Saranya Balachandran and Amarie, \{Oana V.\} and \{Hrab{\v e} de Angelis\}, Martin and Behncke, \{Rose Yinghan\} and Chan, \{Wing Lee\} and Alexandra Despang and Dickel, \{Diane E.\} and Madeleine Duran and Annette Feuchtinger and Helmut Fuchs and Valerie Gailus-Durner and Natja Haag and Rene H{\"a}gerling and Nils Hansmeier and Friederike Hennig and Cooper Marshall and Sudha Rajderkar and Alessa Ringel and Michael Robson and Saunders, \{Lauren M.\} and \{da Silva-Buttkus\}, Patricia and Nadine Spielmann and Srivatsan, \{Sanjay R.\} and Sascha Ulferts and Lars Wittler and Yiwen Zhu and Kalscheuer, \{Vera M.\} and Ibrahim, \{Daniel M.\} and Ingo Kurth and Uwe Kornak and Axel Visel and Pennacchio, \{Len A.\} and Beier, \{David R.\} and Cole Trapnell and Junyue Cao and Jay Shendure and Malte Spielmann",

note = "Publisher Copyright: {\textcopyright} 2023, The Author(s).",

year = "2023",

month = nov,

day = "23",

doi = "10.1038/s41586-023-06548-w",

language = "English",

volume = "623",

pages = "772--781",

journal = "Nature",

issn = "0028-0836",

publisher = "Nature Research",

number = "7988",

}

Huang, X, Henck, J, Qiu, C, Sreenivasan, VKA, Balachandran, S, Amarie, OV, Hrabě de Angelis, M, Behncke, RY, Chan, WL, Despang, A, Dickel, DE, Duran, M, Feuchtinger, A, Fuchs, H, Gailus-Durner, V, Haag, N, Hägerling, R, Hansmeier, N, Hennig, F, Marshall, C, Rajderkar, S, Ringel, A, Robson, M, Saunders, LM, da Silva-Buttkus, P, Spielmann, N, Srivatsan, SR, Ulferts, S, Wittler, L, Zhu, Y, Kalscheuer, VM, Ibrahim, DM, Kurth, I, Kornak, U, Visel, A, Pennacchio, LA, Beier, DR, Trapnell, C, Cao, J, Shendure, J & Spielmann, M 2023, 'Single-cell, whole-embryo phenotyping of mammalian developmental disorders', Nature, vol. 623, no. 7988, pp. 772-781. https://doi.org/10.1038/s41586-023-06548-w

TY - JOUR

T1 - Single-cell, whole-embryo phenotyping of mammalian developmental disorders

AU - Huang, Xingfan

AU - Henck, Jana

AU - Qiu, Chengxiang

AU - Sreenivasan, Varun K.A.

AU - Balachandran, Saranya

AU - Amarie, Oana V.

AU - Hrabě de Angelis, Martin

AU - Behncke, Rose Yinghan

AU - Chan, Wing Lee

AU - Despang, Alexandra

AU - Dickel, Diane E.

AU - Duran, Madeleine

AU - Feuchtinger, Annette

AU - Fuchs, Helmut

AU - Gailus-Durner, Valerie

AU - Haag, Natja

AU - Hägerling, Rene

AU - Hansmeier, Nils

AU - Hennig, Friederike

AU - Marshall, Cooper

AU - Rajderkar, Sudha

AU - Ringel, Alessa

AU - Robson, Michael

AU - Saunders, Lauren M.

AU - da Silva-Buttkus, Patricia

AU - Spielmann, Nadine

AU - Srivatsan, Sanjay R.

AU - Ulferts, Sascha

AU - Wittler, Lars

AU - Zhu, Yiwen

AU - Kalscheuer, Vera M.

AU - Ibrahim, Daniel M.

AU - Kurth, Ingo

AU - Kornak, Uwe

AU - Visel, Axel

AU - Pennacchio, Len A.

AU - Beier, David R.

AU - Trapnell, Cole

AU - Cao, Junyue

AU - Shendure, Jay

AU - Spielmann, Malte

PY - 2023/11/23

Y1 - 2023/11/23

N2 - Mouse models are a critical tool for studying human diseases, particularly developmental disorders1. However, conventional approaches for phenotyping may fail to detect subtle defects throughout the developing mouse2. Here we set out to establish single-cell RNA sequencing of the whole embryo as a scalable platform for the systematic phenotyping of mouse genetic models. We applied combinatorial indexing-based single-cell RNA sequencing3 to profile 101 embryos of 22 mutant and 4 wild-type genotypes at embryonic day 13.5, altogether profiling more than 1.6 million nuclei. The 22 mutants represent a range of anticipated phenotypic severities, from established multisystem disorders to deletions of individual regulatory regions4,5. We developed and applied several analytical frameworks for detecting differences in composition and/or gene expression across 52 cell types or trajectories. Some mutants exhibit changes in dozens of trajectories whereas others exhibit changes in only a few cell types. We also identify differences between widely used wild-type strains, compare phenotyping of gain- versus loss-of-function mutants and characterize deletions of topological associating domain boundaries. Notably, some changes are shared among mutants, suggesting that developmental pleiotropy might be ‘decomposable’ through further scaling of this approach. Overall, our findings show how single-cell profiling of whole embryos can enable the systematic molecular and cellular phenotypic characterization of mouse mutants with unprecedented breadth and resolution.

AB - Mouse models are a critical tool for studying human diseases, particularly developmental disorders1. However, conventional approaches for phenotyping may fail to detect subtle defects throughout the developing mouse2. Here we set out to establish single-cell RNA sequencing of the whole embryo as a scalable platform for the systematic phenotyping of mouse genetic models. We applied combinatorial indexing-based single-cell RNA sequencing3 to profile 101 embryos of 22 mutant and 4 wild-type genotypes at embryonic day 13.5, altogether profiling more than 1.6 million nuclei. The 22 mutants represent a range of anticipated phenotypic severities, from established multisystem disorders to deletions of individual regulatory regions4,5. We developed and applied several analytical frameworks for detecting differences in composition and/or gene expression across 52 cell types or trajectories. Some mutants exhibit changes in dozens of trajectories whereas others exhibit changes in only a few cell types. We also identify differences between widely used wild-type strains, compare phenotyping of gain- versus loss-of-function mutants and characterize deletions of topological associating domain boundaries. Notably, some changes are shared among mutants, suggesting that developmental pleiotropy might be ‘decomposable’ through further scaling of this approach. Overall, our findings show how single-cell profiling of whole embryos can enable the systematic molecular and cellular phenotypic characterization of mouse mutants with unprecedented breadth and resolution.

UR - https://www.scopus.com/pages/publications/85176552712

U2 - 10.1038/s41586-023-06548-w

DO - 10.1038/s41586-023-06548-w

M3 - Journal articles

C2 - 37968388

AN - SCOPUS:85176552712

SN - 0028-0836

VL - 623

SP - 772

EP - 781

JO - Nature

JF - Nature

IS - 7988

ER -

Single-cell, whole-embryo phenotyping of mammalian developmental disorders

Abstract

Funding

UN SDGs

Research Areas and Centers

DFG Research Classification Scheme

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