PI3K和MAPK信号通路对A549细胞FOXO1蛋白表达及其亚细胞定位的调节  

PI3K and MAPK signaling pathways regulate the expression and localization of FOXO1 in A549 cell

作　　者：张洪开[1] 具英花[1] 徐韬钧[1] 孙秀华[1] 于爱鸣[1] 

机构地区：[1]中国医科大学基础医学院生物化学与分子生物学教研室,辽宁沈阳110001

出　　处：《解剖科学进展》2012年第2期126-129,133,共5页Progress of Anatomical Sciences

基　　金：辽宁省教育厅科研基金资助项目(No.2004D225)

摘　　要：目的探讨非小细胞肺癌中磷脂酰肌醇-3-激酶(PI3K)/AKT和丝裂原活化蛋白激酶MAPK/ERK信号通路对Forkhead转录因子(Fox蛋白家族)FOXO1活性的影响及分子机制。方法体外培养人非小细胞肺癌系A549,分别选用PI3K/AKT信号通路特异性抑制剂LY294002和MAPK/ERK信号通路特异性抑制剂UO126及两种抑制剂联合处理细胞之后,MTT比色法检测其对细胞增殖的影响;Western blot检测蛋白FOXO1、p-FOXO1及信号下游蛋白Bim表达的变化;免疫荧光法检测FOXO1蛋白在A549细胞中亚细胞的定位的变化。结果与对照组相比,LY294002和UO126都能明显地抑制A549细胞的增殖,且具有时间依赖性。Western blot结果显示,FOXO1的蛋白水平未见显著性变化,而FOXO1的磷酸化水平明显下降,Bim的表达相应增加。免疫荧光结果显示,FOXO1在A549细胞中的核转位增多。LY294002和UO126联合处理A549细胞时,较药物单独作用效果明显增加。结论 PI3K/AKT和MAPK/ERK信号通路能调节FOXO1磷酸化水平,且具有协同作用,抑制其转录活性。FOXO1通过激活Bim蛋白,促进细胞凋亡,抑制细胞的增殖。Objective To investigate the regulatory mechanism of PI3K and MAPK signaling pathways on FOXO1 transcription factor activity.Methods Non-small cell lung cancer（NSCLC） cell line A549 was cultured in vitro and treated with specific PI3K inhibitor LY294002 and MAPK inhibitor UO126 synergistically/additively.MTT colorimetric assay was employed to plot the cell growth curves,western blot method was used to detect the expression of FOXO1,p-FOXO1 and Bim.Immunofluorescence（IF） analysis was used to examine the localization of FOXO1.Results The growth of A549 cells was siginificantly inhibited by LY294002 and UO126 in a time-dependant manner.Westerm blot revealed that phosphorylation of FOXO1 remarkably reduced and expression of Bim increased by LY294002 and UO126,however,no change was found for the total FOXO1,and FOXO1 was found to be redistributed to the nucleus.The combined effect induced by LY294002 and UO126 was higher than that by LY294002 and UO126 respectively.Conclusion PI3K and MAPK pathways activate phosphorylation of FOXO1,and inhibit FOXO1 transcriptional activity,FOXO1 upregulates the expression of Bim leading cells to apoptosis.

关 键 词：FOXO1 PI3K MAPK 肺癌 

分 类 号：R734.2[医药卫生—肿瘤] Q78[医药卫生—临床医学]