USP7调节EGR1的表达对喉癌细胞生物学行为的影响

Ubiquitin specific protease 7 affects the biological behavior and mechanism of laryngeal cancer cells by regulating the expression of EGR1

摘要:

目的 本研究主要通过siRNA敲低喉癌细胞USP7表达后，检测经转录组测序筛选并与喉癌增殖、迁移相关基因早期生长反应基因1（EGR1）的表达，以了解其在喉癌发病机制中的作用。方法 设计特异性siRNA序列，包装慢病毒并转染喉癌HEp-2细胞特异性敲低泛素特异性蛋白酶7（USP7）表达，再通过RT-PCR和Western blot方法从基因和蛋白质水平检测EGR1的表达。结果 ①siRNA敲低喉癌细胞中USP7后，敲低组USP7 mRNA表达显著降低，同时敲低组USP7蛋白也显著下降；②RT-PCR检测到敲低后的喉癌细胞EGR1 mRNA表达上升，Western blot检测到EGR1蛋白质表达也呈上升；③CCK8实验检测到敲低后的喉癌细胞较阴性对照组和空白组细胞增殖能力明显下降；Transwell实验检测敲低组喉癌细胞迁移能力较阴性对照组和空白组明显下降。结论 在USP7敲低后，EGR1在基因和蛋白水平上升，表明在喉癌的发生过程中USP7可能对EGR1的表达有抑制作用，因此，USP7可能通过抑制EGR1表达在喉癌发生发展过程中有一定作用，为喉癌的临床研究提供实验依据。

关键词:喉癌;泛素特异性蛋白酶7;早期生长反应基因1;立即早期基因

Abstract:

Objective To detect the expression of the early growth response 1 (EGR1) gene after knocked down the expression of ubiquitin specific protease 7 (USP7) of laryngeal cancer cells, so as to understand the pathogenesis of laryngeal carcinoma. Methods Specific siRNA sequences were designed with lentivirus and the speciality of laryngeal carcinoma HEp-2 cells was transfected to knock down USP7 expression, and then the expressions of immediate early (EGR1) gene from the mRNA and protein levels were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Results ①The USP7 mRNA and protein expressions significantly decreased in the knockdown group.② In the knockdown group, RT-PCR revealed increased expression of EGR1 mRNA and Western blot showed elevated expression of EGR1 protein in laryngeal carcinoma cells. ③CCK8 colorimetery revealed that the cell proliferation ability of the knockdown group was significantly lower than those of the negative and blank control groups. Transwell migration assays showed that cell migration ability of the knockdown group was significantly lower than those of the negative and blank control groups. Conclusion Increased expressions of EGR1 at the levels of mRNA and protein after knocking down the USP7 indicates that USP7 may inhibit the expression of EGR1 during the development of laryngeal carcinoma. Therefore, USP7 may play a role in the development of laryngeal cancer by inhibiting the expression of EGR1, providing an experimental evidence for clinical research of laryngeal cancer.

Key words:Laryngeal cancer;Ubiquitin specific protease 7(USP7);Early growth response 1(EGR1);Immediate early gene