目的：探究泛素特异性蛋白酶21（USP21）在胆管癌（CCA）中的表达及其对CCA细胞增殖与迁移的作用及其机制。 方法：通过生物信息学方法和免疫组化及 WB 法检测 CCA 组织及细胞中 USP21 的表达情况。利用体外克隆形成、EdU 及 Transwell实验检测敲低USP21对CCA细胞QBC939和RBE增殖及迁移的影响。通过RNA测序、质谱、免疫共沉淀（Co-IP）及 WB 法探究 USP21 的促癌机制。结果：TCGA 等数据库分析结果显示，USP21 mRNA 在 CCA 组织中呈高表达（均 P < 0.05）。 USP21蛋白在CCA组织和细胞中呈高表达（P < 0.05或P < 0.001或P < 0.000 1）。敲低USP21后，QBC939和RBE细胞的增殖和 迁移能力均显著降低（P < 0.01或P < 0.001）。RNA测序结果表明，敲低USP21可以通过抑制PI3K/AKT信号通路抑制CCA细胞 的增殖和迁移能力（P < 0.05）。质谱鉴定发现，USP21与胰岛素样生长因子2 mRNA结合蛋白1（IGF2BP1）相结合。Co-IP和WB 实验结果表明，USP21与IGF2BP1结合并通过泛素化途径调控IGF2BP1的蛋白表达（P < 0.001或P < 0.000 1）。结论：USP21在 CCA组织和细胞中均呈高表达，其通过IGF2BP1/PI3K/AKT信号通路增强CCA细胞的增殖及迁移能力。

Objectives: To investigate the expression of ubiquitin-specific protease 21 (USP21) in cholangiocarcinoma (CCA) and its effect on the proliferation and migration of CCA cells, as well as the underlying mechanisms. Methods: The expression of USP21 in CCA tissues and cells was detected using bioinformatics approaches, immunohistochemistry, and Western blotting (WB). The effects of USP21 knockdown on the proliferation and migration of CCA cell lines (QBC939 and RBE) were assessed by in vitro colony formation, EdU, and Transwell assays. The oncogenic mechanism of USP21 was explored using RNA sequencing, mass spectrometry, co-immunoprecipitation (CO-IP), and WB. Results: Analysis of databases such as TCGA revealed that USP21 mRNA expression was significantly elevated in CCA tissues (P < 0.05) and USP21 protein expression was highly expressed in both CCA tissues and cells (P < 0.05, P < 0.01, or P < 0.001). Knockdown of USP21 led to a significant decrease in the proliferative and migratory abilities of QBC939 and RBE cells (P < 0.01 or P < 0.001). RNA sequencing results indicated that USP21 knockdown inhibited the proliferative and migratory abilities of CCA cells by suppressing the PI3K/AKT signaling pathway (P < 0.05). Mass spectrometry identified a binding between USP21 and insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1). Co-IP and WB results demonstrated that USP21 interacts with IGF2BP1 and regulates its protein expression through the ubiquitination pathway (P < 0.001 or P < 0.000 1). Conclusion: USP21 is highly expressed in both CCA tissues and cells and enhances the proliferative and migratory abilities of CCA through the IGF2BP1/PI3K/AKT signaling pathway.

江苏省基础研究计划自然科学基金-青年基金项目（No. BK20220723）