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Highly multiplexed spatially resolved gene expression profiling of mouse organogenesis

Lohoff, T. and Ghazanfar, S. and Missarova, A. and Koulena, N. and Pierson, N. and Griffiths, J. A. and Bardot, E. S. and Eng, C.-H. L. and Tyser, R. C. V. and Argelaguet, R. and Guibentif, C. and Srinivas, S. and Briscoe, J. and Simons, B. D. and Hadjantonakis, A.-K. and Gottgens, B. and Reik, W. and Nichols, J. and Cai, L. and Marioni, J. C. (2020) Highly multiplexed spatially resolved gene expression profiling of mouse organogenesis. . (Unpublished)

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Transcriptional and epigenetic profiling of single-cells has advanced our knowledge of the molecular bases of gastrulation and early organogenesis. However, current approaches rely on dissociating cells from tissues, thereby losing the crucial spatial context that is necessary for understanding cell and tissue interactions during development. Here, we apply an image-based single-cell transcriptomics method, seqFISH, to simultaneously and precisely detect mRNA molecules for 387 selected target genes in 8-12 somite stage mouse embryo tissue sections. By integrating spatial context and highly multiplexed transcriptional measurements with two single-cell transcriptome atlases we accurately characterize cell types across the embryo and demonstrate how spatially-resolved expression of genes not profiled by seqFISH can be imputed. We use this high-resolution spatial map to characterize fundamental steps in the patterning of the midbrain-hindbrain boundary and the developing gut tube. Our spatial atlas uncovers axes of resolution that are not apparent from single-cell RNA sequencing data - for example, in the gut tube we observe early dorsal-ventral separation of esophageal and tracheal progenitor populations. In sum, by computationally integrating high-resolution spatially-resolved gene expression maps with single-cell genomics data, we provide a powerful new approach for studying how and when cell fate decisions are made during early mammalian development.

Item Type:Report or Paper (Discussion Paper)
Related URLs:
URLURL TypeDescription Paper ItemData ItemScripts for downstream analysis
Lohoff, T.0000-0001-9333-842X
Ghazanfar, S.0000-0001-7861-6997
Missarova, A.0000-0001-9472-2095
Koulena, N.0000-0002-9419-5712
Pierson, N.0000-0002-2451-0633
Griffiths, J. A.0000-0002-2010-2296
Bardot, E. S.0000-0002-0872-4957
Argelaguet, R.0000-0003-3199-3722
Guibentif, C.0000-0001-8457-456X
Srinivas, S.0000-0001-5726-7791
Briscoe, J.0000-0002-1020-5240
Simons, B. D.0000-0002-3875-7071
Gottgens, B.0000-0001-6302-5705
Reik, W.0000-0003-0216-9881
Cai, L.0000-0002-7154-5361
Additional Information:The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license. This version posted November 21, 2020. We thank our colleagues in the Wellcome Trust Mouse Gastrulation Consortium, as well as colleagues in the University of Cambridge Stem Cell Institute, Cancer Research UK Cambridge Institute, Babraham Institute, Gurdon Institute, California Institute of Technology, Sloan Kettering, and the Francis Crick Institute for their support and intellectual engagement. We thank Julian Thomassie, Elsy Buitrago-Delgado, Jina Yun, Michael Lawson, Christopher Cronin, Chris Karp and other members of the Cai Lab for experimental support and constructive input. We thank Sonja Nowotschin and Esther Wershof for discussions concerning the gut tube analysis. We thank Veronique Juvin for providing scientific illustration. We thank Daniel Keitley and Michael Morgan and other members of the Marioni Lab for discussions concerning the analysis. We thank members of the Nichols and Reik lab for their discussions concerning the project. Data availability: The spatial transcriptomic map can be explored interactively at: and raw image data is available on request. Processed gene expression data with segmentation information and associated metadata is also available to download and explore online at Scripts for downstream analysis are available at Author Contributions: T.L., J.A.G. and C.G., performed the probe library gene selection, with input from W.R., J.N., B.G. and J.C.M.. T.L., E.S.B. and J.N. performed embryo collection. T.L., C.-H.L.E and N.K. performed the seqFISH method and segmentation optimization for mouse embryonic tissue sections. T.L. and N.K. generated the spatial dataset, with supervision from L.C.. N.P. performed image processing including registration, cell segmentation and mRNA spot decoding, with input from L.C.. T.L. and A.M. performed optical threshold selection for non-barcoded smFISH images. S.G. performed pre-processing, low-level analyses, batch correction, clustering, integration with other datasets, global visualization, and designed the associated website, with input from A.M., R.A.. A.M. performed imputation analysis. S.G. and A.M. performed analysis surrounding midbrain hindbrain boundary formation. E.S.B. performed HCR imaging experiments, with supervision from A.-K.H.. R.T., C.G., S.S., J.B., B.D.S., A.-K.H., B.G., W.R. and J.N. provided discussion and interpretation of the data and analysis. B.G., W.R., J.N., L.C., and J.C.M. supervised the study. T.L., S.G. and A.M. generated figures. T.L., S.G., A.M. and J.C.M. wrote the manuscript. L.C. and J.C.M. oversaw the entirety of the project. All authors read and approved the final manuscript. The following sources of funding are gratefully acknowledged. This work was supported by the Wellcome Trust (award 105031/D/14/Z) to W.R, J.N., J.C.M., B.G., S.S., and B.D.S.. T.L. was was funded by the Wellcome Trust 4-Year PhD Programme in Stem Cell Biology and Medicine and the University of Cambridge, UK (203813/Z/16/A; 203813/Z/16/Z) and Boehringer Ingelheim Fonds travel grant. S.G. was supported by a Royal Society Newton International Fellowship (NIF\R1\181950). A.M. was supported by an NIH award (1OT2OD026673-01 – Comprehensive Collaborative, Infrastructure, Mapping and Tools for the HubMAP HIVE (Mapping Component) to J.C.M.). C.G. was supported by funding from the Swedish Research Council (2017-06278). A.-K.H. was supported by National Institutes of Health (NIH) grants (award numbers R01-DK084391 and P30-CA008748). B.G. and J.N. are supported by core funding by the MRC and Wellcome Trust to the Wellcome–MRC Cambridge Stem Cell Institute. L.C. was supported by the Paul G. Allen Frontiers Foundation Discovery Center for Cell Lineage Tracing (grant UWSC10142). J.C.M. acknowledges core funding from EMBL and core support from Cancer Research UK (C9545/A29580). The funding sources mentioned above had no role in the study design; in the collection, analysis, and interpretation of data, in the writing of the manuscript, and in the decision to submit the manuscript for publication. Competing Interest Statement: W.R. is a consultant and shareholder of Cambridge Epigenetix. L.C. is the co-founder of Spatial Genomics Inc. and holds patents on seqFISH.
Funding AgencyGrant Number
Wellcome Trust105031/D/14/Z
Wellcome Trust203813/Z/16/A
Wellcome Trust203813/Z/16/Z
Boehringer Ingelheim FondsUNSPECIFIED
Royal SocietyNIF\R1\181950
Swedish Research Council2017-06278
Medical Research Council (UK)UNSPECIFIED
Paul G. Allen Frontiers GroupUWSC10142
European Molecular Biology Laboratory (EMBL)UNSPECIFIED
Cancer Research UKC9545/A29580
Record Number:CaltechAUTHORS:20201123-133350121
Persistent URL:
Official Citation:Highly multiplexed spatially resolved gene expression profiling of mouse organogenesis. Tim Lohoff, Shila Ghazanfar, Alsu Missarova, Noushin Koulena, Nico Pierson, Jonathan A. Griffiths, Evan S. Bardot, Chee-Huat Linus Eng, Richard C. V. Tyser, Ricard Argelaguet, Carolina Guibentif, Shankar Srinivas, James Briscoe, Benjamin David Simons, Anna-Katerina Hadjantonakis, Berthold Gottgens, Wolf Reik, Jenny Nichols, Long Cai, John C. Marioni. bioRxiv 2020.11.20.391896; doi:
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:106790
Deposited By: Tony Diaz
Deposited On:23 Nov 2020 22:16
Last Modified:30 Jan 2021 00:50

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