p53在苯并[a]芘诱导所致p21和细胞周期蛋白改变中的作用  

Roles of p53 in the interaction of p21 and cell cycle proteins induced by benzo[a]pyrene

作　　者：张夏男[1] 李跃纲[2] 贾效伟[1] 刘秉慈[1] 叶萌[1] 

机构地区：[1]中国疾病预防控制中心职业卫生与中毒控制所化学污染物与健康重点实验室,北京100050 [2]北京市疾病预防控制中心营养与食品卫生所

出　　处：《卫生研究》2014年第2期193-197,共5页Journal of Hygiene Research

基　　金：国家自然科学基金(No.30972449;30671747)

摘　　要：目的探讨在苯并[a]芘(B[a]P)诱导的人胚肺成纤维细胞(HELF)中p53蛋白对p21、cyclin D1和CDK4蛋白间相互作用的影响。方法向转染p53小干扰RNA(p53siRNA)质粒的HELF细胞即p53-H细胞组、载体CMV的HELF细胞即HELF/CMV细胞组中分别加入2μmol/L B[a]P作用24 h,向p53化学抑制剂Pifithrin-α(PFT-α)组即HELF/CMV+PFT-α细胞组中同时加入2μmol/L B[a]P和20μmol/L PFT-α作用24 h,各组同时设立不做任何处理的对照组。用免疫印迹(Western blot)法检测细胞中p53、p53-ser20以及p21、cyclin D1和CDK4蛋白水平的变化,同时利用免疫沉淀方法分析p53蛋白对p21、cyclin D1以及CDK4蛋白间相互作用的影响。结果利用PFT-α或p53 siRNA技术抑制p53蛋白后,B[a]P诱导的p53蛋白20位丝氨酸磷酸化水平和p21蛋白水平的增高受抑制,B[a]P诱导的cyclin D1水平的增加不受影响,CDK4的水平不受B[a]P的影响;免疫沉淀实验结果表明,B[a]P引起的p21和CDK4结合的增加受到抑制,B[a]P引起的p21与cyclin D1结合的增加不受影响,cyclin D1和CDK4的结合不受B[a]P的影响。结论B[a]P通过p53-ser20影响人胚肺成纤维细胞p21和CDK4的结合。Objective To investigate the roles of p53 in the interaction of p21, cyclin D1 and CDK4 in human embryonic lung fibroblasts （HELFs） induced by benzo （a） pyrene. Methods p53-H group （cells transfected with p53 small interference RNA plasmid, p53 siRNA） and HELF/CMV group （cells transfected with CMV vector） were treated with 2 txmol/L B [ a]P for 24 h, and HELF/CMV ＋ PFT-α group （HELF/CMV cells were treated with p53 chemical inhibitor, Pifithrin-α） was treated with 2 ixmol/L B [a]P and 20 μmol/L PFT-α for 24 h. The above three groups set up control groups, respectively. Western blot assay was used to check the levels of p53, phosphorylated p53 at 20 site （p53-ser20）, p21, cyclin D1 and CDK4. Immunopreeipitation assay was used to investigate the roles of p53 in the interaction of p21, cyclin D1 and CDK4. Results After inhibition of p53 using PFT-α or siRNA, the high levels of p53, p53-ser 20 and p2! induced by B [ a]P were markedly decreased. The change of cyelin D1 level was not obsevered and the level CDK4 was free of B[ a]P. The combination of p21 and CDK 4 was increased after HELFs exposed to B [ a] P, which can not be observed after inhibition p53. The combination of p21 and cyelin D1 was increased with or without the expression of p53 after HELFs exposed to B[a]P. The combination of eyclin D1 and CDK 4 was not affected by B [ a]P. Conclusion p53 can affect the combination of p21 and CDK4 in HELFs induced by B[ a]P.

关 键 词：苯并[A]芘 P53 细胞周期调控蛋白 免疫沉淀 

分 类 号：R994.6[医药卫生—毒理学] R730.231[医药卫生—药学]