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. 2024 Jan 15;9:18. doi: 10.1038/s41392-023-01709-9

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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 — Bim level was increased with impaired ubiquitination degradation in DN T cells for neddylation inhibition. a The proteins annotated with apoptosis were selected and ranked based on the ratio value from the proteomic results of DN T cells. Red dots represented proteins significantly upregulated (FC＞ 1.5, P < 0.05); Blue dots represented proteins significantly downregulated (FC < 0.667, P < 0.05). n = 4/group. b, c The mRNA levels of Ube2m and Bim in DN T cells were evaluated via real-time PCR. n = 3/group. *P < 0.05, **P < 0.01. d Immunoblotting assay and densitometric analysis evaluated the protein level of Bim in DN T cells treated with MG-132. *P < 0.05. e Western blot assay was conducted to determine the level of Cullin1 neddylation, Ube2m and Bim in DN T cells. One band represented one mouse. n = 3. f Ubiquitination degradation of Bim in DN T cells was measured via Co-IP assay. Data were representative of three independent experiments. g The apoptosis of DN T cells treated with MLN4924 were detected via flow cytometry (Annexin V⁺/7-AAD^-). h The apoptosis of DN T cells treated with MLN4924 was quantified based on the analysis of flow cytometry. n = 3/group. *P < 0.05. i Immunoblotting assay and densitometric analysis showed the level of Bim and Cleaved-caspase3 in MLN4924 treated-DN T cells. *P < 0.05. j Ubiquitination degradation of Bim in MLN4924-treated DN T cells was measured via Co-IP assay. Data were representative of three independent experiments