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. 2023 Sep 21;30(11):1640–1652. doi: 10.1038/s41594-023-01096-3

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© The Author(s) 2023

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. 5 — a, Overview of the Hic1^CreERT2 knock-in allele⁵⁰ and of the Rosa26-hSSM2 allele (Rosa26^hSS2⁾⁵¹ for conditional induction of SS18-SSX2 in Hic1-expressing mesenchymal progenitors. Upon tamoxifen treatment, CreERT2 mediates recombination between the two LoxP sites in SSM2 mice, thereby removing the transcriptional stop signal and allowing transcription of SS18-SSX2⁻IRES-EGFP from the endogenous ROSA26 promoter. b, Illustration of the timeline for the tissue sample collection of samples analyzed in c and d. Eight-week-old mice were treated with tamoxifen, and tongue muscle tissues were collected at 5, 7 and 9 weeks after induction. c,d, Immunofluorescence of Hic1^{creERT2/creERT2}; Rosa26^SSM2/SSM2, Cre-positive mouse tongue tissue at 5, 7 or 9 weeks after induction. The cells are stained for DAPI, SSM2 (eGFP) and H2AK119ub1 (c) or BCOR (d). Scale bars, 100 μm. e, Quantification of H2AK119ub1 (top) and BCOR (bottom) signal intensity normalized to DAPI signal intensity in three biological replicates (three different mice) in tamoxifen treated mice (+TAM) expressing or not expressing the SSM2 cassette (human SS18-SSX2) and showing normal tongue muscle (+TAM; SSM2⁻) adjacent to synovial sarcoma tumors (+TAM; SSM2⁺). P values determined by paired one-tailed t-test between groups (from left to right, *P = 0.04, *P = 0.04, **P = 0.002 (H2AK119ub1); *P = 0.03, *P = 0.04, *P = 0.04 (BCOR)). f, Immunohistochemical staining for H2AK119ub1 on a tissue microarray of human surgical excised tissue specimens (left, skeletal muscle; right, synovial sarcoma). Scale bar, 50 μm. g, Quantification of H2AK119ub DAB signal intensity across 37 synovial sarcomas (sample cores in duplicate), other sarcomas (one case each of epithelioid sarcoma, sarcomatoid mesothelioma, Ewing sarcoma, sarcomatoid renal cell carcinoma, clear cell sarcoma, dedifferentiated liposarcoma and myxoid liposarcoma) and normal tissues (normal skeletal muscle, ovarian stroma, breast glandular tissue and testis controls). Quantification for the two skeletal muscle samples is also shown separately in the graph. All samples were stained in parallel on the same formalin-fixed, paraffin-embedded tissue microarray slide. P values determined by Mann–Whitney U-test between groups (**P = 0.001 (H2AK119ub1), ****P < 0.0001 (SS18-SSX)).

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