High-resolution transcriptional and morphogenetic profiling of cells from micropatterned human ESC gastruloid cultures
- Washington University in St Louis, United States
- Washington University School of Medicine, United States
- University of California, Davis, United States
- University of Illinois, Urbana-Champaign, United States
Abstract
During mammalian gastrulation, germ layers arise and are shaped into the body plan while extraembryonic layers sustain the embryo. Human embryonic stem cells, cultured with BMP4 on extracellular matrix micro-discs, reproducibly differentiate into gastruloids, expressing markers of germ layers and extraembryonic cells in radial arrangement. Using single-cell RNA sequencing and cross-species comparisons with mouse, cynomolgus monkey gastrulae, and post-implantation human embryos, we reveal that gastruloids contain cells transcriptionally similar to epiblast, ectoderm, mesoderm, endoderm, primordial germ cells, trophectoderm, and amnion. Upon gastruloid dissociation, single cells reseeded onto micro-discs were motile and aggregated with the same but segregated from distinct cell types. Ectodermal cells segregated from endodermal and extraembryonic but mixed with mesodermal cells. Our work demonstrates that the gastruloid system models primate-specific features of embryogenesis, and that gastruloid cells exhibit evolutionarily conserved sorting behaviors. This work generates a resource for transcriptomes of human extraembryonic and embryonic germ layers differentiated in a stereotyped arrangement.
Data availability
Sequencing data have been deposited in GEO under accession code GSE144897.
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In vitro culture of cynomolgus monkey embryos beyond gastrulationNCBI Gene Expression Omnibus, GSE130114.
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A developmental coordinate of the spectrum of pluripotency among mice, monkeys, and humansNCBI Gene Expression Omnibus, GSE74767.
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A developmental landscape of 3D-cultured human pre-gastrulation embryosNCBI Gene Expression Omnibus, GSE136447.
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Human Primordial Germ Cells are Specified from Lineage Primed ProgenitorsNCBI Gene Expression Omnibus, GSE140021.
Article and author information
Author details
Lilianna Solnica-Krezel
Funding
Children's Discovery Institute
- Lilianna Solnica-Krezel
Washington University School of Medicine in St. Louis (Discretionary funds)
- Lilianna Solnica-Krezel
Vallee Foundation (Vallee Scholar Award)
- Samantha A Morris
Paul G. Allen Frontiers Group (Allen Distinguished Investigator Award)
- Samantha A Morris
Alfred P. Sloan Foundation (Sloan Research Fellowship)
- Samantha A Morris
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Anita Bhattacharyya, University of Wisconsin, Madison, United States
Version history
- Received: May 28, 2020
- Accepted: November 17, 2020
- Accepted Manuscript published: November 18, 2020 (version 1)
- Version of Record published: December 10, 2020 (version 2)
Copyright
© 2020, Minn et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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High-resolution transcriptional and morphogenetic profiling of cells from micropatterned human ESC gastruloid cultures
eLife 9:e59445.
https://doi.org/10.7554/eLife.59445
Further reading
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From a forward mutagenetic screen to discover mutations associated with obesity, we identified mutations in the Spag7 gene linked to metabolic dysfunction in mice. Here, we show that SPAG7 KO mice are born smaller and develop obesity and glucose intolerance in adulthood. This obesity does not stem from hyperphagia, but a decrease in energy expenditure. The KO animals also display reduced exercise tolerance and muscle function due to impaired mitochondrial function. Furthermore, SPAG7-deficiency in developing embryos leads to intrauterine growth restriction, brought on by placental insufficiency, likely due to abnormal development of the placental junctional zone. This insufficiency leads to loss of SPAG7-deficient fetuses in utero and reduced birth weights of those that survive. We hypothesize that a ‘thrifty phenotype’ is ingrained in SPAG7 KO animals during development that leads to adult obesity. Collectively, these results indicate that SPAG7 is essential for embryonic development and energy homeostasis later in life.
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- Developmental Biology
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Amniogenesis, a process critical for continuation of healthy pregnancy, is triggered in a collection of pluripotent epiblast cells as the human embryo implants. Previous studies have established that bone morphogenetic protein (BMP) signaling is a major driver of this lineage specifying process, but the downstream BMP-dependent transcriptional networks that lead to successful amniogenesis remain to be identified. This is, in part, due to the current lack of a robust and reproducible model system that enables mechanistic investigations exclusively into amniogenesis. Here, we developed an improved model of early amnion specification, using a human pluripotent stem cell-based platform in which the activation of BMP signaling is controlled and synchronous. Uniform amniogenesis is seen within 48 hr after BMP activation, and the resulting cells share transcriptomic characteristics with amnion cells of a gastrulating human embryo. Using detailed time-course transcriptomic analyses, we established a previously uncharacterized BMP-dependent amniotic transcriptional cascade, and identified markers that represent five distinct stages of amnion fate specification; the expression of selected markers was validated in early post-implantation macaque embryos. Moreover, a cohort of factors that could potentially control specific stages of amniogenesis was identified, including the transcription factor TFAP2A. Functionally, we determined that, once amniogenesis is triggered by the BMP pathway, TFAP2A controls the progression of amniogenesis. This work presents a temporally resolved transcriptomic resource for several previously uncharacterized amniogenesis states and demonstrates a critical intermediate role for TFAP2A during amnion fate specification.
