scMultiome analysis identifies embryonic hindbrain progenitors with mixed rhombomere identities

  1. Yong-Il Kim
  2. Rebecca O'Rourke
  3. Charles G Sagerström  Is a corresponding author
  1. University of Colorado Anschutz Medical Campus, United States

Abstract

Rhombomeres serve to position neural progenitors in the embryonic hindbrain, thereby ensuring appropriate neural circuit formation, but the molecular identities of individual rhombomeres and the mechanism whereby they form have not been fully established. Here we apply scMultiome analysis in zebrafish to molecularly resolve all rhombomeres for the first time. We find that rhombomeres become molecularly distinct between 10hpf (end of gastrulation) and 13hpf (early segmentation). While the embryonic hindbrain transiently contains alternating odd- versus even-type rhombomeres, our scMultiome analyses do not detect extensive odd versus even molecular characteristics in the early hindbrain. Instead, we find that each rhombomere displays a unique gene expression and chromatin profile. Prior to the appearance of distinct rhombomeres, we detect three hindbrain progenitor clusters (PHPDs) that correlate with the earliest visually observed segments in the hindbrain primordium and that represent prospective rhombomere r2/r3 (possibly including r1), r4 and r5/r6, respectively. We further find that the PHPDs form in response to Fgf and RA morphogens and that individual PHPD cells co-express markers of multiple mature rhombomeres. We propose that the PHPDs contain mixed-identity progenitors and that their subdivision into individual rhombomeres requires resolution of mixed transcription and chromatin states.

Data availability

scMultiome data has been deposited at GEO under record number GSE223535

The following data sets were generated

Article and author information

Author details

  1. Yong-Il Kim

    Department of Pediatrics, University of Colorado Anschutz Medical Campus, Aurora, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Rebecca O'Rourke

    Department of Pediatrics, University of Colorado Anschutz Medical Campus, Aurora, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1198-6963
  3. Charles G Sagerström

    Department of Pediatrics, University of Colorado Anschutz Medical Campus, Aurora, United States
    For correspondence
    charles.sagerstrom@cuanschutz.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1509-5810

Funding

National Institute of Neurological Disorders and Stroke (NS038183)

  • Charles G Sagerström

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: The Institutional Animal Care and Use Committee (IACUC) of the University of Colorado Medical School approved all procedures involving zebrafish under protocol #870.

Copyright

© 2023, Kim 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.

Metrics

  • 897
    views
  • 116
    downloads
  • 1
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Yong-Il Kim
  2. Rebecca O'Rourke
  3. Charles G Sagerström
(2023)
scMultiome analysis identifies embryonic hindbrain progenitors with mixed rhombomere identities
eLife 12:e87772.
https://doi.org/10.7554/eLife.87772

Share this article

https://doi.org/10.7554/eLife.87772

Further reading

    1. Cancer Biology
    2. Developmental Biology
    Sara Jaber, Eliana Eldawra ... Franck Toledo
    Research Article

    Missense ‘hotspot’ mutations localized in six p53 codons account for 20% of TP53 mutations in human cancers. Hotspot p53 mutants have lost the tumor suppressive functions of the wildtype protein, but whether and how they may gain additional functions promoting tumorigenesis remain controversial. Here, we generated Trp53Y217C, a mouse model of the human hotspot mutant TP53Y220C. DNA damage responses were lost in Trp53Y217C/Y217C (Trp53YC/YC) cells, and Trp53YC/YC fibroblasts exhibited increased chromosome instability compared to Trp53-/- cells. Furthermore, Trp53YC/YC male mice died earlier than Trp53-/- males, with more aggressive thymic lymphomas. This correlated with an increased expression of inflammation-related genes in Trp53YC/YC thymic cells compared to Trp53-/- cells. Surprisingly, we recovered only one Trp53YC/YC female for 22 Trp53YC/YC males at weaning, a skewed distribution explained by a high frequency of Trp53YC/YC female embryos with exencephaly and the death of most Trp53YC/YC female neonates. Strikingly, however, when we treated pregnant females with the anti-inflammatory drug supformin (LCC-12), we observed a fivefold increase in the proportion of viable Trp53YC/YC weaned females in their progeny. Together, these data suggest that the p53Y217C mutation not only abrogates wildtype p53 functions but also promotes inflammation, with oncogenic effects in males and teratogenic effects in females.

    1. Developmental Biology
    Mengjie Li, Aiguo Tian, Jin Jiang
    Research Advance

    Stem cell self-renewal often relies on asymmetric fate determination governed by niche signals and/or cell-intrinsic factors but how these regulatory mechanisms cooperate to promote asymmetric fate decision remains poorly understood. In adult Drosophila midgut, asymmetric Notch (N) signaling inhibits intestinal stem cell (ISC) self-renewal by promoting ISC differentiation into enteroblast (EB). We have previously shown that epithelium-derived Bone Morphogenetic Protein (BMP) promotes ISC self-renewal by antagonizing N pathway activity (Tian and Jiang, 2014). Here, we show that loss of BMP signaling results in ectopic N pathway activity even when the N ligand Delta (Dl) is depleted, and that the N inhibitor Numb acts in parallel with BMP signaling to ensure a robust ISC self-renewal program. Although Numb is asymmetrically segregated in about 80% of dividing ISCs, its activity is largely dispensable for ISC fate determination under normal homeostasis. However, Numb becomes crucial for ISC self-renewal when BMP signaling is compromised. Whereas neither Mad RNA interference nor its hypomorphic mutation led to ISC loss, inactivation of Numb in these backgrounds resulted in stem cell loss due to precocious ISC-to-EB differentiation. Furthermore, we find that numb mutations resulted in stem cell loss during midgut regeneration in response to epithelial damage that causes fluctuation in BMP pathway activity, suggesting that the asymmetrical segregation of Numb into the future ISC may provide a fail-save mechanism for ISC self-renewal by offsetting BMP pathway fluctuation, which is important for ISC maintenance in regenerative guts.