Theranostics 2019; 9(2):573-587. doi:10.7150/thno.27654

Research Paper

USP5 promotes epithelial-mesenchymal transition by stabilizing SLUG in hepatocellular carcinoma

Jing Meng1*, Xiaoyu Ai1*, Yueyang Lei1*, Weilong Zhong1*, Baoxin Qian3, Kailiang Qiao1, Xiaorui Wang4, Bijiao Zhou1, Hongzhi Wang1, Longcong Huai1, Xiaoyun Zhang1, Jingxia Han2, Yinyin Xue1, Yuan Liang1, Honggang Zhou1,2✉, Shuang Chen2✉, Tao Sun1,2✉, Cheng Yang1,2✉

1. State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Nankai University, Tianjin, China
2. Tianjin Key Laboratory of Molecular Drug Research, Tianjin International Joint Academy of Biomedicine, Tianjin, China
3. Department of Gastroenterology and Hepatology, Tianjin Key Laboratory of Artificial Cells, Tianjin Institute of Hepatobiliary Disease, Tianjin Third Central Hospital, Tianjin, China
4. College of Life Science, Nankai University, Tianjin, China
* These authors contributed equally to this work.


Rationale: The role of SLUG in epithelial-mesenchymal transition during tumor progression has been thoroughly studied, but its precise regulation remains poorly explored.

Methods: The affinity purification, mass spectrometry and CO-IP were performed to identify the interaction between SLUG and ubiquitin-specific protease 5 (USP5). Cycloheximide chase assays and deubiquitination assays confirmed that the effect of USP5 on the deubiquitin of SLUG. The dual-luciferase reporter and chromatin immunoprecipitation assays were employed to observe the direct transcriptional regulation of E-cadherin by SLUG effected by USP5. EMT related markers was detected by western blotting and immunofluorescence. Molecular docking, SPR sensor (biacore) and co-location were detected to prove Formononetin targets USP5.

Bioinformatics analysis was used to study the relation of USP5 and SLUG to malignancy degree of HCC. Cell migration, invasion in HCC cells and xenografts model in nude mouse were conducted to detect the promotion of USP5 and the inhibition of Formononetin on EMT.

Results: USP5 interacts with and stabilizes SLUG to regulate its abundance through USP5 deubiquitination activities in epithelial-mesenchymal transition (EMT) of hepatocellular carcinoma (HCC). USP5 is highly expressed and positively correlated with SLUG expression in HCC with high malignancy. Knockdown of USP5 inhibits SLUG deubiquitination and inhibits HCC cells proliferation, metastasis, and invasion, while overexpression of USP5 promotes SLUG stability and EMT in vitro and in vivo. Through virtual screening, we found that Formononetin exhibits excellent binding to USP5. Moreover, Formononetin inhibits deubiquitinating activities of USP5 to SLUG and consequently impedes the EMT and malignant progression of HCC.

Conclusion: Our findings reveal that USP5 serve as a potential target for tumor intervention and provide a preliminary antitumor therapy for inhibit EMT by targeting USP5 or its interaction with SLUG in HCC.

Keywords: USP5, SLUG, EMT, Formononetin, hepatocellular carcinoma

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How to cite this article:
Meng J, Ai X, Lei Y, Zhong W, Qian B, Qiao K, Wang X, Zhou B, Wang H, Huai L, Zhang X, Han J, Xue Y, Liang Y, Zhou H, Chen S, Sun T, Yang C. USP5 promotes epithelial-mesenchymal transition by stabilizing SLUG in hepatocellular carcinoma. Theranostics 2019; 9(2):573-587. doi:10.7150/thno.27654. Available from