TRIM21通过泛素化稳定细胞骨架蛋白质促进下咽癌分化

TRIM21 Promotes Differentiation of Hypopharyngeal Squamous Cell Carcinoma Through Improving Protein Stability by Ubiquitination of Cytoskeleton

E3泛素化连接酶TRIM21(tripartite motif containing 21)在不同类型肿瘤中扮演癌基因或抑癌基因的角色,从而影响细胞功能及临床预后。然而,TRIM21在下咽癌中的生物学功能和分子机制尚不清楚。本文采用RT-PCR和Western印迹技术检测了TRIM21在下咽癌组织中的表达水平,采用基因过表达和RNA干扰技术分析了TRIM21过表达和基因沉默对细胞增殖、迁移和分化的影响,利用蛋白质免疫共沉淀联合质谱鉴定TRIM21互作蛋白质,并进一步研究TRIM21影响下咽癌分化的分子机制。结果表明,TRIM21在中高分化下咽癌中高表达,提示TRIM21可能调控肿瘤细胞分化。在FaDu细胞中,过表达或shRNA干扰TRIM21可分别抑制和促进细胞增殖和迁移。此外,过表达TRIM21后,分化标志分子角蛋白10(keratin 10,KRT10)、内披蛋白(involucrin,IVL)和谷氨酰胺转氨酶1(transglutaminase 1,TGM1)蛋白质表达水平增高,而shRNA干扰TRIM21后,上述蛋白质表达下降。质谱鉴定及蛋白质生物信息分析提示,TRIM21可能与细胞骨架蛋白质调控密切相关。进一步验证表明,TRIM21与角蛋白10相互作用。蛋白质合成抑制剂环己酰亚胺处理细胞后,与对照相比,TRIM21过表达细胞中角蛋白10表达增高,且过表达TRIM21可升高角蛋白10的泛素化水平。综上所述,我们的研究表明,TRIM21可能通过介导细胞骨架蛋白质泛素化修饰,提高蛋白质稳定性促进下咽癌分化。

E3 ubiquitin ligase TRIM21(tripartite motif containing 21)plays an important role in regulating cell biological functions and clinical prognosis as oncogene or tumor suppressor in different types of tumors.However,the biological functions and molecular mechanism of TRIM21 in hypopharyngeal squamous cell carcinoma(HPSCC)are still unclear.Our results showed that TRIM21 is highly expressed in moderately and well differentiated HPSCC,suggesting the role of TRIM21 in tumor differentiation.Overexpression and knockdown of TRIM21 inhibited or promoted cell proliferation and migration.Meanwhile,the expression of differentiation markers including KRT10(keratin 10),IVL(involucrin)and TGM1(transglutaminase 1)were increased and decreased upon TRIM21 overexpression or knockdown,respectively.The bioinformatics analysis of TRIM21 interacting protein identified by co-immunoprecipitation combined with mass spectrometry suggested that TRIM21 may be closely related to the regulation of cytoskeleton protein.We further demonstrated that TRIM21 interacted with KRT10.The inhibition of protein synthesis by cycloheximide led to upregulation of KRT10 in TRIM21 overexpressing FaDu cells,and ectopic expression of TRIM21 enhanced the ubiquitination level of KRT10.In summary,our results suggest that TRIM21 may promote the HPSCC differentiation by mediating ubiquitination of cytoskeleton proteins to improve the protein stability.