Combined transient ablation and single cell RNA sequencing reveals the development of medullary thymic epithelial cells

  1. Kristen L Wells
  2. Corey N Miller
  3. Andreas R Gschwind
  4. Wu Wei
  5. Jonah D Phipps
  6. Mark S Anderson  Is a corresponding author
  7. Lars M Steinmetz  Is a corresponding author
  1. Stanford University, United States
  2. University of California, San Francisco, United States
  3. European Molecular Biology Laboratory, Germany

Abstract

Medullary thymic epithelial cells (mTECs) play a critical role in central immune tolerance by mediating negative selection of autoreactive T cells through the collective expression of the peripheral self-antigen compartment, including tissue-specific antigens (TSAs). Recent work has shown that gene expression patterns within the mTEC compartment are remarkably heterogenous and include multiple differentiated cell states. To further define mTEC development and medullary epithelial lineage relationships, we combined lineage tracing and recovery from transient in vivo mTEC ablation with single cell RNA-sequencing in Mus musculus. The combination of bioinformatic and experimental approaches revealed a non-stem transit-amplifying population of cycling mTECs that preceded Aire expression. Based on our findings, we propose a branching model of mTEC development wherein a heterogeneous pool of transit-amplifying cells gives rise to Aire- and Ccl21a-expressing mTEC subsets. We further use experimental techniques to show that within the Aire-expressing developmental branch, TSA expression peaked as Aire expression decreased, implying Aire expression must be established before TSA expression can occur. Collectively, these data provide a higher order roadmap of mTEC development and demonstrate the power of combinatorial approaches leveraging both in vivo models and high-dimensional datasets.

Data availability

RNA-seq data that support the findings of this study have been deposited in the GEO database under accession numbers GSE137699

The following data sets were generated

Article and author information

Author details

  1. Kristen L Wells

    Genetics, Stanford University, Stanford, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7466-8164
  2. Corey N Miller

    Diabetes Center, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Andreas R Gschwind

    Genetics, Stanford University, Stanford, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0769-6907
  4. Wu Wei

    Genetics, Stanford University, Stanford, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Jonah D Phipps

    Diabetes Center, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Mark S Anderson

    Diabetes Center, University of California, San Francisco, San Francisco, United States
    For correspondence
    Mark.Anderson@ucsf.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3093-4758
  7. Lars M Steinmetz

    Genome Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany
    For correspondence
    lars.steinmetz@embl.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3962-2865

Funding

National Science Foundation (DGE- 1656518)

  • Kristen L Wells

National Institutes of Health (P01 HG000205)

  • Lars M Steinmetz

National Institutes of Health (R01 AI097457)

  • Mark S Anderson

National Institutes of Health (R01 AI097457)

  • Mark S Anderson

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

Ethics

Animal experimentation: Mice were maintained in the University of California San Francisco (UCSF) specific pathogen- free animal facility in accordance with the guidelines established by the Institutional Animal Care and Use Committee (IACUC) and Laboratory Animal Resource Center and all experimental procedures were approved by the Laboratory Animal Resource Center at UCSF. The animal protocol number associated with the study is AN180637-02B.

Reviewing Editor

  1. Ellen A Robey, University of California, Berkeley, United States

Publication history

  1. Received: June 18, 2020
  2. Accepted: November 21, 2020
  3. Accepted Manuscript published: November 23, 2020 (version 1)
  4. Version of Record published: December 29, 2020 (version 2)

Copyright

© 2020, Wells 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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  1. Kristen L Wells
  2. Corey N Miller
  3. Andreas R Gschwind
  4. Wu Wei
  5. Jonah D Phipps
  6. Mark S Anderson
  7. Lars M Steinmetz
(2020)
Combined transient ablation and single cell RNA sequencing reveals the development of medullary thymic epithelial cells
eLife 9:e60188.
https://doi.org/10.7554/eLife.60188

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