# Regulation of medullary thymic epithelial cells and thymic central tolerance by Ikaros

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2024 · $675,743

## Abstract

Project Summary
A normal and robust immune system relies on the development of cells with diverse repertoire of antigen
recognition but that remain tolerant of self-tissues. Breakdown of this immune tolerance can give rise to
autoimmune disease, and hence multiple mechanisms are in place to ensure immune self-tolerance. The thymus
is a critical site for the development and education of T cells to promote tolerance to self through expression of
tissue specific antigens (TSAs) by specialized medullary thymic epithelial cells (mTECs). The transcriptional
regulators Aire and Fezf2 are known to act within mTECs to promote the expression of thousands of TSA self-
antigens for the purpose of removing developing self-reactive T cells in a process known as negative selection.
Aire and Fezf2 are shown to be required for some, but not all TSA genes expressed in mTECs, suggesting
additional transcriptional regulators are required for the full repertoire of the observed TSA expression in mTECs.
Furthermore, recent work has highlighted the complexity of mTECs with at least four mTEC populations: Aire+
mTECs, Ccl21a mTECs, Late/Post Aire mTECs, and tuft cells. However, there is limited knowledge about the
developmental regulation and progenitors of these heterogeneous mTEC subsets. We have identified the
transcription factor Ikaros (Ikzf1) as a novel regulator of mTEC composition and function, with deletion of Ikaros
in mTECs causing a reduction of Aire+ mTECs and an expansion of tuft cells. Moreover, Ikaros-deficient mTECs
have a defect in TSA gene expression resulting in specific signs of autoimmunity. Interestingly, mutations in
IKZF1 have been linked to human autoimmune diseases. Thus, we hypothesize a novel role for Ikaros in
mTEC lineage development, TSA expression and central tolerance, and that defects in mTEC function could
contribute to the autoimmunity seen in humans with IKZF1 mutations. We propose to test our hypothesis and
increase our knowledge of this novel mTEC transcriptional regulator through the following specific aims: (1)
Identify the stage(s) of mTEC development at which Ikaros modulates mTEC function, (2) Interrogate the
molecular mechanism of Ikaros function in mTECs, and (3) Determine if TEC specific deletion of Ikaros affects
T cell tolerance and autoimmunity. To complete these aims, we have developed a unique and powerful set of
genetic tools and mouse reporter lines that allow us to perturb Ikaros function in mTECs. We will use flow
cytometry to investigate mTEC and thymic immune cell phenotypes, combined scRNA-seq/scATAC-seq to
analyze gene expression and chromatin accessibility, and supplement with mechanistic studies in vitro in cell
lines. Our long-term goal is to increase our understanding of the regulation of mTEC development and function
in central tolerance, and how alterations can lead to a break of tolerance and autoimmunity.

## Key facts

- **NIH application ID:** 10753566
- **Project number:** 5R01AI165829-02
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** Michael R Waterfield
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $675,743
- **Award type:** 5
- **Project period:** 2022-12-12 → 2027-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10753566

## Citation

> US National Institutes of Health, RePORTER application 10753566, Regulation of medullary thymic epithelial cells and thymic central tolerance by Ikaros (5R01AI165829-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10753566. Licensed CC0.

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