Hedgehog regulation of epithelial cell state and morphogenesis in the larynx

  1. Janani Ramachandran
  2. Weiqiang Zhou
  3. Anna E Bardenhagen
  4. Talia Nasr
  5. Ellen R Yates
  6. Aaron M Zorn
  7. Hongkai Ji
  8. Steven A Vokes  Is a corresponding author
  1. The University of Texas at Austin, United States
  2. Johns Hopkins University, United States
  3. Cincinnati Children's Hospital Medical Center, United States

Abstract

The larynx enables speech while regulating swallowing and respiration. Larynx function hinges on the laryngeal epithelium which originates as part of the anterior foregut and undergoes extensive remodeling to separate from the esophagus and form vocal folds that interface with the adjacent trachea. Here we find that Sonic hedgehog (SHH) is essential for epithelial integrity in the mouse larynx as well as the anterior foregut. During larynx-esophageal separation, low Shh expression marks specific domains of actively remodeling epithelium that undergo an epithelial to mesenchymal transition (EMT) characterized by the induction of N-Cadherin and movement of cells out of the epithelial layer. Consistent with a role for SHH signaling in regulating this process, Shh mutants undergo an abnormal EMT throughout the anterior foregut and larynx, marked by a cadherin switch, movement out of the epithelial layer and cell death. Unexpectedly, Shh mutant epithelial cells are replaced by a new population of FOXA2-negative cells that likely derive from adjacent pouch tissues and form a rudimentary epithelium. These findings have important implications for interpreting the etiology of HH-dependent birth defects within the foregut. We propose that SHH signaling has a default role in maintaining epithelial identity throughout the anterior foregut and that regionalized reductions in SHH trigger epithelial remodeling.

Data availability

Sequencing data have been deposited in GEO under accession code GSE190281.

The following data sets were generated

Article and author information

Author details

  1. Janani Ramachandran

    Department of Molecular Biosciences, The University of Texas at Austin, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Weiqiang Zhou

    Department of Biostatistics, Johns Hopkins University, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Anna E Bardenhagen

    Department of Molecular Biosciences, The University of Texas at Austin, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Talia Nasr

    Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2473-5402
  5. Ellen R Yates

    Department of Molecular Biosciences, The University of Texas at Austin, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Aaron M Zorn

    Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3217-3590
  7. Hongkai Ji

    Department of Biostatistics, Johns Hopkins University, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Steven A Vokes

    Department of Molecular Biosciences, The University of Texas at Austin, Austin, United States
    For correspondence
    svokes@austin.utexas.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1724-0102

Funding

National Institutes of Health (RO1 HD090163)

  • Hongkai Ji
  • Steven A Vokes

National Institutes of Health (RO1 HD093363)

  • Aaron M Zorn

National Institutes of Health (F30 HL142201)

  • Talia Nasr

University of Texas at Austin (Continuing Graduate Fellowship)

  • Janani Ramachandran

University of Texas at Austin (Provost's Graduate Excellence Fellowship)

  • Janani Ramachandran

University of Texas at Austin (TIDES Summer Fellowship)

  • Anna E Bardenhagen

University of Texas at Austin (Experiential Learning Summer Scholarship)

  • Ellen R Yates

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

Reviewing Editor

  1. Marianne E Bronner, California Institute of Technology, United States

Ethics

Animal experimentation: All experiments involving mice were approved by the Institutional Animal Care and Use Committee at the University of Texas at Austin (protocol AUP-2019-00233).

Version history

  1. Received: January 14, 2022
  2. Preprint posted: January 20, 2022 (view preprint)
  3. Accepted: November 18, 2022
  4. Accepted Manuscript published: November 18, 2022 (version 1)
  5. Version of Record published: December 2, 2022 (version 2)

Copyright

© 2022, Ramachandran 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

  • 695
    views
  • 117
    downloads
  • 3
    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. Janani Ramachandran
  2. Weiqiang Zhou
  3. Anna E Bardenhagen
  4. Talia Nasr
  5. Ellen R Yates
  6. Aaron M Zorn
  7. Hongkai Ji
  8. Steven A Vokes
(2022)
Hedgehog regulation of epithelial cell state and morphogenesis in the larynx
eLife 11:e77055.
https://doi.org/10.7554/eLife.77055

Share this article

https://doi.org/10.7554/eLife.77055

Further reading

    1. Chromosomes and Gene Expression
    2. Developmental Biology
    F Javier DeHaro-Arbona, Charalambos Roussos ... Sarah Bray
    Research Article

    Developmental programming involves the accurate conversion of signalling levels and dynamics to transcriptional outputs. The transcriptional relay in the Notch pathway relies on nuclear complexes containing the co-activator Mastermind (Mam). By tracking these complexes in real time, we reveal that they promote the formation of a dynamic transcription hub in Notch ON nuclei which concentrates key factors including the Mediator CDK module. The composition of the hub is labile and persists after Notch withdrawal conferring a memory that enables rapid reformation. Surprisingly, only a third of Notch ON hubs progress to a state with nascent transcription, which correlates with polymerase II and core Mediator recruitment. This probability is increased by a second signal. The discovery that target-gene transcription is probabilistic has far-reaching implications because it implies that stochastic differences in Notch pathway output can arise downstream of receptor activation.

    1. Developmental Biology
    Rieko Asai, Vivek N Prakash ... Takashi Mikawa
    Research Article

    Large-scale cell flow characterizes gastrulation in animal development. In amniote gastrulation, particularly in avian gastrula, a bilateral vortex-like counter-rotating cell flow, called ‘polonaise movements’, appears along the midline. Here, through experimental manipulations, we addressed relationships between the polonaise movements and morphogenesis of the primitive streak, the earliest midline structure in amniotes. Suppression of the Wnt/planar cell polarity (PCP) signaling pathway maintains the polonaise movements along a deformed primitive streak. Mitotic arrest leads to diminished extension and development of the primitive streak and maintains the early phase of the polonaise movements. Ectopically induced Vg1, an axis-inducing morphogen, generates the polonaise movements, aligned to the induced midline, but disturbs the stereotypical cell flow pattern at the authentic midline. Despite the altered cell flow, induction and extension of the primitive streak are preserved along both authentic and induced midlines. Finally, we show that ectopic axis-inducing morphogen, Vg1, is capable of initiating the polonaise movements without concomitant PS extension under mitotic arrest conditions. These results are consistent with a model wherein primitive streak morphogenesis is required for the maintenance of the polonaise movements, but the polonaise movements are not necessarily responsible for primitive streak morphogenesis. Our data describe a previously undefined relationship between the large-scale cell flow and midline morphogenesis in gastrulation.