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. 2023 Oct 26;11:56. doi: 10.1038/s41413-023-00296-3

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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. 4 — YAP1/β-catenin signaling is essential for Trim21-mediated osteogenic differentiation. a Schematic diagram showing TMT-based quantitative proteomics for the identification of differentially expressed proteins (DEPs) in bone marrow mesenchymal stem cells (BMSCs) derived from Trim21^+/+ and Trim21^−/− mice. b Volcano plots of DEPs in BMSCs from Trim21^+/+ and Trim21^−/− mice. c Heatmap analysis of DEPs in BMSCs. Three replicates of each group were included, and the top 29 DEPs are shown. d KEGG enrichment analysis of the DEPs in the BMSCs. e Representative immunoblotting analysis and quantification of BCL9, AXIN1, and YAP1 in BMSCs; proteomics sample: part of the samples subjected to proteomics analysis. f Representative immunoblotting analysis and quantification of BCL9, β-catenin, YAP1, and Runx2 protein expression in BMSCs after osteogenic induction for 7 days. g The endogenous interaction between Trim21, BCL9, β-catenin, and YAP1 was evaluated using a co-IP assay. h Protein‒protein interaction of YAP1 and Trim21 in living cells. The two BiFC plasmids encoding Myc-VN155-YAP1 and HA-VC155-Trim21 along with HA-cerulean were cotransfected into HEK293T cells for 24 h. Representative images showing transfected cells (cerulean) and the interaction between YAP1 and Trim21 (Venus). Nuclei were stained with DAPI. Scale bar: 20 μm. i Immunoblotting analysis of BCL9, β-catenin, YAP1, and HA-Trim21 protein expression in HEK293T cells treated with or without MG132. j Representative images showing the expression of YAP1⁺ OBs derived from Trim21^+/+ and Trim21^−/− mice. Scale bar: 20 μm. All bar graphs are presented as the mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.000 1; n.s. not significant by Student’s t test