泛素蛋白连接酶E3A在乳腺癌细胞中作用的蛋白质组学与生物信息学分析

Proteomics and bioinformatics analyses of role of ubiquitin-protein ligase E3A in breast cancer cells

目的:通过蛋白质组学与生物信息学方法探讨泛素蛋白连接酶E3A(UBE3A)在乳腺癌细胞中的生物学功能。方法:将三阴性乳腺癌细胞MDA-MB-231分别转染shRNA-UBE3A片段(UBE3A敲低组)与阴性对照shRNA片段(对照组)后,采用双向电泳(2-DE)法及基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-TOF-MS/MS)对两组细胞的差异蛋白质分离和鉴定;采用DAVID Functional Annotation及String在线工具对差异表达蛋白质进行生物信息学分析。结果:2-DE及MALDI-TOF-TOF-MS/MS分离和鉴定出28个差异蛋白,其中UBE3A敲低组相对于对照组有4个表达上调,24个表达下调。与DAVID数据库有23个相匹配,其中能够进行生物学途径、细胞成分、分子功能注释分类的蛋白质分别为23个(100%)、20个(87.7%)和23个(100%),而在泛素-蛋白酶体系统及糖代谢方面分类比较明确;String分析显示这些差异蛋白中有23个存在直接或间接的联系。结论:UBE3A可能通过泛素-蛋白酶体系统功能与糖代谢途径参与乳腺癌细胞生物学行为的调控。

Objective: To investigate the potential biological function of ubiquitin-protein ligase E3A (UBE3A) in breast cancer cells through proteomics and bioinformatics methods. Methods: The triple-negative breast cancer MDA-MB-231 cells were transfected with shRNA-UBE3A sequences (UBE3A knock-down group) or scrambled shRNA sequences (control group), respectively. Then, the differentially expressed proteins between the two groups of cells were separated and identified by two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionization (MALDI-TOF-TOF-MS/MS);bioinformatics analyses of the differentially expressed proteins were performed using DAVID Functional Annotation and String online tools. Results: Twenty-eight differentially expressed proteins were obtained and identified by 2-DE and MALDI-TOF-TOF-MS/MS, and 23 proteins were matched in DAVID database, in which, 4 were up-regulated, 24 were down-regulated in UBE3A knock-down group versus control group. Twenty-three proteins were recognized by DAVID database, in which, 23 (100%), 20 (87.7%) and 23 (100%) proteins could be annotated by biological pathway, cellular components and annotation of molecule functions especially, some of them were clearly related to the ubiquitin-proteasome system and glucose metabolism. String analysis showed that there were direct or indirect relations among the 23 proteins. Conclusion: UBE3A may participate in regulation of the biological behaviors of breast cancer cells via the ubiquitin-proteasome system and glucose metabolism pathway.