Recording morphogen signals reveals mechanisms underlying gastruloid symmetry breaking.

Publisher: Macmillan Magazines Ltd Country of Publication: England NLM ID: 100890575 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1476-4679 (Electronic) Linking ISSN: 14657392 NLM ISO Abbreviation: Nat Cell Biol Subsets: MEDLINE

Original Publication: London : Macmillan Magazines Ltd., [1999-

Body Patterning*/genetics ; Gene Expression Regulation, Developmental*; Animals ; Wnt Signaling Pathway ; Signal Transduction ; Wnt Proteins/metabolism ; Wnt Proteins/genetics ; Organoids/metabolism ; Gastrula/metabolism ; Nodal Protein/metabolism ; Nodal Protein/genetics ; Stem Cells/metabolism ; Gene Regulatory Networks ; Cell Aggregation

Aggregates of stem cells can break symmetry and self-organize into embryo-like structures with complex morphologies and gene expression patterns. Mechanisms including reaction-diffusion Turing patterns and cell sorting have been proposed to explain symmetry breaking but distinguishing between these candidate mechanisms of self-organization requires identifying which early asymmetries evolve into subsequent tissue patterns and cell fates. Here we use synthetic 'signal-recording' gene circuits to trace the evolution of signalling patterns in gastruloids, three-dimensional stem cell aggregates that form an anterior-posterior axis and structures resembling the mammalian primitive streak and tailbud. We find that cell sorting rearranges patchy domains of Wnt activity into a single pole that defines the gastruloid anterior-posterior axis. We also trace the emergence of Wnt domains to earlier heterogeneity in Nodal activity even before Wnt activity is detectable. Our study defines a mechanism through which aggregates of stem cells can form a patterning axis even in the absence of external spatial cues.
Competing Interests: Competing interests J.E.T. is a scientific advisor for Prolific Machines and Nereid Therapeutics. B.A. is an advisory board member with options for Arbor Biotechnologies and Tessera Therapeutics, and holds equity in Celsius Therapeutics. H.M.M. is a cofounder and scientific advisor for C16 Biosciences. The remaining authors declare no conflicts of interest.
(© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Update of: bioRxiv. 2023 Jun 05:2023.06.02.543474. doi: 10.1101/2023.06.02.543474. (PMID: 37333235)

Bardot, E. S. & Hadjantonakis, A.-K. Mouse gastrulation: coordination of tissue patterning, specification and diversification of cell fate. Mech. Dev. 163, 103617 (2020). (PMID: 32473204753458510.1016/j.mod.2020.103617)
Gregor, T., Wieschaus, E. F., McGregor, A. P., Bialek, W. & Tank, D. W. Stability and nuclear dynamics of the bicoid morphogen gradient. Cell 130, 141–152 (2007). (PMID: 17632061225367210.1016/j.cell.2007.05.026)
Lord, N. D., Carte, A. N., Abitua, P. B. & Schier, A. F. The pattern of nodal morphogen signaling is shaped by co-receptor expression. eLife 10, e54894 (2021). (PMID: 34036935826638910.7554/eLife.54894)
Turner, D. A. et al. Anteroposterior polarity and elongation in the absence of extra-embryonic tissues and of spatially localised signalling in gastruloids: mammalian embryonic organoids. Development 144, 3894–3906 (2017). (PMID: 289514355702072)
Beccari, L. et al. Multi-axial self-organization properties of mouse embryonic stem cells into gastruloids. Nature 562, 272–276 (2018). (PMID: 3028313410.1038/s41586-018-0578-0)
Veenvliet, J. V. et al. Mouse embryonic stem cells self-organize into trunk-like structures with neural tube and somites. Science 370, eaba4937 (2020). (PMID: 3330358710.1126/science.aba4937)
Miao, Y. et al. Reconstruction and deconstruction of human somitogenesis in vitro. Nature 614, 500–508 (2023). (PMID: 3654332110.1038/s41586-022-05655-4)
Ishihara, K. & Tanaka, E. M. Spontaneous symmetry breaking and pattern formation of organoids. Curr. Opin. Syst. Biol. 11, 123–128 (2018). (PMID: 10.1016/j.coisb.2018.06.002)
van den Brink, S. C. et al. Symmetry breaking, germ layer specification and axial organisation in aggregates of mouse embryonic stem cells. Development 141, 4231–4242 (2014). (PMID: 25371360430291510.1242/dev.113001)
Scoones, J. C., Banerjee, D. S. & Banerjee, S. Size-regulated symmetry breaking in reaction-diffusion models of developmental transitions. Cells 9, 1646 (2020). (PMID: 10.3390/cells9071646)
Sozen, B., Cornwall-Scoones, J. & Zernicka-Goetz, M. The dynamics of morphogenesis in stem cell-based embryology: novel insights for symmetry breaking. Dev. Biol. 474, 82–90 (2021). (PMID: 3333306710.1016/j.ydbio.2020.12.005)
Simunovic, M. et al. A 3D model of a human epiblast reveals BMP4-driven symmetry breaking. Nat. Cell Biol. 21, 900–910 (2019). (PMID: 3126326910.1038/s41556-019-0349-7)
Sick, S., Reinker, S., Timmer, J. & Schlake, T. WNT and DKK determine hair follicle spacing through a reaction-diffusion mechanism. Science 314, 1447–1450 (2006). (PMID: 1708242110.1126/science.1130088)
Raspopovic, J., Marcon, L., Russo, L. & Sharpe, J. Digit patterning is controlled by a Bmp–Sox9–Wnt Turing network modulated by morphogen gradients. Science 345, 566–570 (2014). (PMID: 2508270310.1126/science.1252960)
Glover, J. D. et al. The developmental basis of fingerprint pattern formation and variation. Cell 186, 940–956 (2023). (PMID: 3676429110.1016/j.cell.2023.01.015)
Hashmi, A. et al. Cell-state transitions and collective cell movement generate an endoderm-like region in gastruloids. eLife 11, e59371 (2022). (PMID: 35404233903330010.7554/eLife.59371)
Srivastava, V. et al. Configurational entropy is an intrinsic driver of tissue structural heterogeneity. Preprint at bioRxiv https://doi.org/10.1101/2023.07.01.546933 (2023).
Shyer, A. E. et al. Emergent cellular self-organization and mechanosensation initiate follicle pattern in the avian skin. Science 357, 811–815 (2017). (PMID: 28705989560527710.1126/science.aai7868)
Palmquist, K. H. et al. Reciprocal cell–ECM dynamics generate supracellular fluidity underlying spontaneous follicle patterning. Cell 185, 1960–1973 (2022). (PMID: 3555176510.1016/j.cell.2022.04.023)
Kondo, S., Watanabe, M. & Miyazawa, S. Studies of Turing pattern formation in zebrafish skin. Philos. Trans. R. Soc. A 379, 20200274 (2021). (PMID: 10.1098/rsta.2020.0274)
Korinek, V. et al. Constitutive transcriptional activation by a β-catenin–Tcf complex in APC ^−/− colon carcinoma. Science 275, 1784–1787 (1997). (PMID: 906540110.1126/science.275.5307.1784)
Suppinger, S. et al. Multimodal characterization of murine gastruloid development. Cell Stem Cell https://doi.org/10.1016/j.stem.2023.04.018 (2023).
Serup, P. et al. Partial promoter substitutions generating transcriptional sentinels of diverse signaling pathways in embryonic stem cells and mice. Dis. Model. Mech. 5, 956–966 (2012). (PMID: 228880973484877)
Morgani, S. M., Metzger, J. J., Nichols, J., Siggia, E. D. & Hadjantonakis, A.-K. Micropattern differentiation of mouse pluripotent stem cells recapitulates embryo regionalized cell fate patterning. eLife 7, e32839 (2018). (PMID: 29412136580705110.7554/eLife.32839)
Grosswendt, S. et al. Epigenetic regulator function through mouse gastrulation. Nature 584, 102–108 (2020). (PMID: 32728215741573210.1038/s41586-020-2552-x)
van den Brink, S. C. et al. Single-cell and spatial transcriptomics reveal somitogenesis in gastruloids. Nature 582, 405–409 (2020). (PMID: 3207626310.1038/s41586-020-2024-3)
Merle, M., Friedman, L., Chureau, C., Shoushtarizadeh, A. & Gregor, T. Precise and scalable self-organization in mammalian pseudo-embryos. Nat. Struct. Mol. Biol. 31, 896–902 (2024).
Underhill, E. J. & Toettcher, J. E. Control of gastruloid patterning and morphogenesis by the Erk and Akt signaling pathways. Development 150, dev201663 (2023). (PMID: 375901311110666710.1242/dev.201663)
Anand, G. M. et al. Controlling organoid symmetry breaking uncovers an excitable system underlying human axial elongation. Cell 186, 497–512 (2023). (PMID: 366574431012250910.1016/j.cell.2022.12.043)
Chal, J. & Pourquié, O. Making muscle: skeletal myogenesis in vivo and in vitro. Development 144, 2104–2122 (2017). (PMID: 2863427010.1242/dev.151035)
Wang, X. et al. The development of highly potent inhibitors for porcupine. J. Med. Chem. 56, 2700–2704 (2013). (PMID: 23477365363127410.1021/jm400159c)
Steinberg, M. S. On the mechanism of tissue reconstruction by dissociated cells. I. Population kinetics, differential adhesiveness, and the absence of directed migration. Proc. Natl Acad. Sci. USA 48, 1577–1582 (1962). (PMID: 1391668922100210.1073/pnas.48.9.1577)
Toda, S., Blauch, L. R., Tang, S. K. Y., Morsut, L. & Lim, W. A. Programming self-organizing multicellular structures with synthetic cell–cell signaling. Science 361, 156–162 (2018). (PMID: 29853554649294410.1126/science.aat0271)
Stevens, A. J. et al. Programming multicellular assembly with synthetic cell adhesion molecules. Nature 614, 144–152 (2023). (PMID: 3650910710.1038/s41586-022-05622-z)
Tsai, T. Y.-C. et al. An adhesion code ensures robust pattern formation during tissue morphogenesis. Science 370, 113–116 (2020). (PMID: 33004519787947910.1126/science.aba6637)
Redmer, T. et al. E‐cadherin is crucial for embryonic stem cell pluripotency and can replace OCT4 during somatic cell reprogramming. EMBO Rep. 12, 720–726 (2011). (PMID: 21617704312897110.1038/embor.2011.88)
Soncin, F. et al. E-cadherin acts as a regulator of transcripts associated with a wide range of cellular processes in mouse embryonic stem cells. PLoS ONE 6, e21463 (2011). (PMID: 21779327313647110.1371/journal.pone.0021463)
Xu, P.-F., Houssin, N., Ferri-Lagneau, K. F., Thisse, B. & Thisse, C. Construction of a vertebrate embryo from two opposing morphogen gradients. Science 344, 87–89 (2014). (PMID: 2470085710.1126/science.1248252)
Massey, J. et al. Synergy with TGFβ ligands switches WNT pathway dynamics from transient to sustained during human pluripotent cell differentiation. Proc. Natl Acad. Sci. USA 116, 4989–4998 (2019). (PMID: 30819898642146510.1073/pnas.1815363116)
Camacho-Aguilar, E., Yoon, S., Ortiz-Salazar, M. A. & Warmflash, A. Combinatorial interpretation of BMP and WNT controls the decision between primitive streak and extraembryonic fates. Cell Syst. 15, 445–461 (2024). (PMID: 3869227410.1016/j.cels.2024.04.001)
Mayran, A. et al. Cadherins modulate the self-organizing potential of gastruloids. Preprint at bioRxiv https://doi.org/10.1101/2023.11.22.568291 (2023).
Dingare, C., Cao, D., Yang, J. J., Sozen, B. & Steventon, B. Mannose controls mesoderm specification and symmetry breaking in mouse gastruloids. Dev. Cell 59, 1523–1537 (2024).
Arias, A. M., Marikawa, Y. & Moris, N. Gastruloids: pluripotent stem cell models of mammalian gastrulation and embryo engineering. Dev. Biol. 488, 35–46 (2022). (PMID: 35537519947718510.1016/j.ydbio.2022.05.002)
Pinheiro, D., Kardos, R., Hannezo, É. & Heisenberg, C.-P. Morphogen gradient orchestrates pattern-preserving tissue morphogenesis via motility-driven unjamming. Nat. Phys. 18, 1482–1493 (2022). (PMID: 10.1038/s41567-022-01787-6)
Schüle, K. M. et al. Eomes restricts Brachyury functions at the onset of mouse gastrulation. Dev. Cell 58, 1627–1642 (2023). (PMID: 3763327110.1016/j.devcel.2023.07.023)
Anlas, K. & Trivedi, V. Studying evolution of the primary body axis in vivo and in vitro. eLife 10, e69066 (2021). (PMID: 34463611845673910.7554/eLife.69066)
Petersen, C. P. & Reddien, P. W. Wnt signaling and the polarity of the primary body axis. Cell 139, 1056–1068 (2009). (PMID: 2000580110.1016/j.cell.2009.11.035)
Petersen, C. P. & Reddien, P. W. Polarized notum activation at wounds inhibits Wnt function to promote planarian head regeneration. Science 332, 852–855 (2011). (PMID: 21566195332072310.1126/science.1202143)
Feinberg, S., Roure, A., Piron, J. & Darras, S. Antero–posterior ectoderm patterning by canonical Wnt signaling during ascidian development. PLoS Genet. 15, e1008054 (2019). (PMID: 30925162645757210.1371/journal.pgen.1008054)
Wang, R., Steele, R. E. & Collins, E.-M. S. Wnt signaling determines body axis polarity in regenerating Hydra tissue fragments. Dev. Biol. 467, 88–94 (2020). (PMID: 3287115610.1016/j.ydbio.2020.08.012)
Lovas, J. R. & Yuste, R. Dissociation and reaggregation of Hydra vulgaris for studies of self-organization. STAR Protoc. 3, 101504 (2022). (PMID: 36042889942053210.1016/j.xpro.2022.101504)
Ferenc, J. et al. Mechanical oscillations orchestrate axial patterning through Wnt activation in Hydra. Sci. Adv. 7, eabj6897 (2021). (PMID: 34890235866425710.1126/sciadv.abj6897)
Amadei, G. et al. Embryo model completes gastrulation to neurulation and organogenesis. Nature 610, 143–153 (2022). (PMID: 36007540953477210.1038/s41586-022-05246-3)
Tarazi, S. et al. Post-gastrulation synthetic embryos generated ex utero from mouse naive ESCs. Cell 185, 3290–3306 (2022). (PMID: 35988542943972110.1016/j.cell.2022.07.028)
Bhattarai-Kline, S. et al. Recording gene expression order in DNA by CRISPR addition of retron barcodes. Nature 608, 217–225 (2022). (PMID: 35896746935718210.1038/s41586-022-04994-6)
Choi, J. et al. A time-resolved, multi-symbol molecular recorder via sequential genome editing. Nature 608, 98–107 (2022). (PMID: 35794474935258110.1038/s41586-022-04922-8)
Thielicke, W. & Sonntag, R. Particle image velocimetry for MATLAB: accuracy and enhanced algorithms in PIVlab. J. Open Res. Softw. 9, 12 (2021).

T32GM007388 U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS); U01 DK127429 United States DK NIDDK NIH HHS; R01 GM144362 United States GM NIGMS NIH HHS; 2134935 NSF | Directorate for Biological Sciences (BIO); 1734030 NSF | Directorate for Mathematical & Physical Sciences | Division of Physics (PHY); U01DK127429 U.S. Department of Health & Human Services | NIH | National Institute of Diabetes and Digestive and Kidney Diseases (National Institute of Diabetes & Digestive & Kidney Diseases)

0 (Wnt Proteins)
0 (Nodal Protein)

Date Created: 20241002 Date Completed: 20241116 Latest Revision: 20241211

20241211

10.1038/s41556-024-01521-9

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