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目的研究肺腺癌中与耐药相关的泛素化或类泛素化蛋白质及其与耐药的关系。方法从经顺铂处理和未处理的A549细胞及其耐顺铂细胞株中分离泛素化、类泛素化蛋白质,通过1D SDS-PAGE和Chip HPLC-MS/MS分析鉴别蛋白,Western blot法验证质谱结果;MTT法检测细胞对顺铂的耐药性。结果鉴定出发生类泛素化修饰蛋白质-PKM2,并发现其在耐药细胞株中低表达;以蛋白酶体抑制剂MG132干预细胞后,PKM2表达上调;CDDP和MG132联合作用细胞后,细胞耐药性下降。结论细胞中PKM2表达水平与肿瘤细胞耐药程度呈负相关,PKM2受蛋白质类泛素化修饰-SUMO化修饰调节,提示PKM2可能与肿瘤耐药的发生密切相关。
OBJECTIVE: To study the drug resistance-associated ubiquitinated or ubiquitinated proteins in lung adenocarcinoma and its relationship with drug resistance. Methods Ubiquitinated and ubiquitinated proteins were isolated from cisplatin-treated and untreated A549 cells and their cisplatin-resistant cell lines. Proteins were identified by 1D SDS-PAGE and Chip HPLC-MS / MS analysis. Western blot Mass spectrometry results were validated; MTT assay was used to detect cell resistance to cisplatin. Results The ubiquitin-modified protein-PKM2 was identified and found to be underexpressed in drug-resistant cell lines; PKM2 was up-regulated after proteasome inhibitor MG132 was interfered; CDDP and MG132 combined with cytosolic Sexual decline. Conclusion The expression of PKM2 in cells is negatively correlated with the drug resistance of tumor cells. PKM2 is regulated by protein ubiquitination-SUMOylation, suggesting that PKM2 may be closely related to the development of drug resistance.