HepG2细胞中PIG11相互作用蛋白质的初步分离与鉴定  

Initial Separation and Identification of the PIG11 Interacting Proteins in Human Hepatocellular Carcinoma HepG2 Cells

作　　者：胡蓉[1] 王晓娟[1] 王燕[1] 梁晓秋[1] 

机构地区：[1]南华大学肿瘤研究所,湖南衡阳421001

出　　处：《南华大学学报（医学版）》2009年第2期177-179,共3页Journal of Nanhua University(Medical Edition)

摘　　要：目的探讨人肝细胞癌细胞系(HepG2)并初步鉴定候选肝癌抑癌蛋白肿瘤蛋白P53下游靶基因11号(PIG11)相互作用蛋白,确定PIG11的作用方式。方法课题组前期利用质粒pLXSN构建逆转录病毒载体,转染细胞后建立起PIG11高表达HepG2细胞株。将HepG2分为HepG2对照组,空载体HepG2组和PIG11高表达HepG2组分别培养,流式细胞术检测细胞凋亡。采用免疫共沉淀富集HepG2细胞中PIG11结合蛋白,切取PIG11结合蛋白条带,胶内酶解后进行电喷雾串联质谱得到肽序列标签,通过数据库查询蛋白。结果PIG11蛋白高表达组凋亡率显著高于对照组(P<0.01)。串联质谱后数据库查询鉴定出11个蛋白,分别是热休克蛋白家族中的HSP60,KH型剪接调节蛋白(KSRP)、脑酸溶性蛋白BASP1、蛋白水解诱导因子(PIF/DCD)、Y染色体RNA结合蛋白、碳酸酐酶、白蛋白以及骨架蛋白中的波形蛋白、α、β-微管蛋白、肌动蛋白。结论PIG11蛋白高表达对HepG2具有诱导凋亡作用;初步鉴定出11个PIG11结合蛋白,为进一步阐明PIG11蛋白的作用机制提供线索。Objective To find out the proteins interacting with tumor protein P53 inducible protein 11 （ PIG11 ） and get more information of the mechanism of PIG11 over - expresssion in human hepatoblastoma cell line （ HepG2 ） cells. Methods The plasmid pLXSN was used to construct the human PIG11 gene retroviral vector, and PIGll gene high expression HepG2 cell line was achieved after transfected. HepG2, pLXSN - HepG2, pLXSN - PIG11 - HepG2 group were cultured, and cell apoptosis was analyzed by flow cytometer scans. PIG11 bingding proteins were recoverd by Co - immunoprecipitation, protein identification was performed by Mass spectrometry, peptide sequence tags were analyzed by Mascot database. Results The apopetosis rate of HepG2 - pLXSN - PIG11 was higher than that of HepG2 - pLXSN and the control HepG2. Mass spectrometry analysis allowed the identification of 11 PIG11 interacting proteins： Hsp60, KSRP, Brain abundant membrane attached Signal protein 1 （ BASP1 ） , Dermcidin （ DCD/PIF ） , Y - chromosome RNA recognition Motif protein, Carbonate dehydratase, ALB protein, Vimentin, Alpha - tubulin, Tubulin 5 - bete and Actin. Conclusion The findings suggest that over - expression of PIG11 induce HepG2 cells apoptosis. 11 PIG11 interacting proteins were identified, which can provide us a clue on further studying the function of PIG11.

关 键 词：PIG11 HEPG2 免疫共沉淀 电喷雾串联质谱 

分 类 号：R735.7[医药卫生—肿瘤]