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. 2022 Sep 7;610(7933):752–760. doi: 10.1038/s41586-022-05309-5

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a, UMAP visualization of integrated 10X Genomics and Smart-seq2 (SS2) scRNA-seq analysis for RORγt⁺MHCII⁺ Thetis cells (TC), coloured by SMART-seq2 Thetis cell transcriptome or 10X cluster annotation. b, Intracellular expression of AIRE protein by Thetis cells and dendritic cells. c, Index-sorting summary graphs for CD11c, CD11b cell-surface protein and RORγt (Venus) fluorescence intensity. d, Heat map showing expression of top differentially expressed genes (DEGs) between Thetis cells and MHCII⁺ ILC3s, profiled by SMART-seq2, identifying Rora as an ILC3–Thetis cell-distinguishing gene. e, Representative flow cytometry analysis of tdTomato expression in MHCII⁺ILC3 and Thetis cells isolated from mLN of Rorc^VenusRora^creRosa26^lsl-tdTomato fate-mapped mice at P14 (n = 4). f, Heat map reporting scaled chromVAR deviation transcription factor motif scores (left) and corresponding transcription factor gene expression values (right) for top transcription factor gene–motif pairs in Thetis cells in scATAC-seq data. g, Heat map showing scaled, imputed expression of top 125 DEGs (one versus the rest, fold change (FC) > 1.5, adjusted P < 0.01) for each Thetis cell cluster. h, Dot plot showing expression of selected cell-surface markers that are differentially expressed between Thetis cell subsets. i, Gating strategy for identification of Thetis cell subsets. j, Intracellular expression of AIRE protein by Thetis cell subsets; each symbol represents an individual mouse (n = 4). k, summary of TC I–TC IV phenotypes. Plots in i are representative of n = 6 mice from 3 independent experiments. Data in b,e,j are representative of 3 independent experiments. Box plots in c indicate median (centre line) and interquartile range (hinges), whiskers represent minimum and maximum values, and dots represent outliers.

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