[1]杨卫文,张月,谢雄,等.纤维粘连蛋白EDA片段抑制P53核转位调控肠癌SW480细胞凋亡[J].第三军医大学学报,2017,39(08):787-793.
 Yang Weiwen,Zhang Yue,Xie Xiong,et al.Fibronectin extra domain A suppresses apoptosis of colorectal cancer SW480 cells by inhibiting P53 nuclear translocation[J].J Third Mil Med Univ,2017,39(08):787-793.
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纤维粘连蛋白EDA片段抑制P53核转位调控肠癌SW480细胞凋亡(/HTML )
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
39卷
期数:
2017年第08期
页码:
787-793
栏目:
基础医学
出版日期:
2017-04-30

文章信息/Info

Title:
Fibronectin extra domain A suppresses apoptosis of colorectal cancer SW480 cells by inhibiting P53 nuclear translocation
作者:
杨卫文张月谢雄欧娟娟梁后杰庞学利
第三军医大学西南医院肿瘤科
Author(s):
Yang Weiwen Zhang Yue Xie Xiong Ou Juanjuan Liang Houjie Pang Xueli

Department of Oncology, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China

关键词:
纤维粘连蛋白EDA片段P53凋亡结直肠癌
Keywords:
fibronectin extra domain A P53 apoptosis colorectal cancer
分类号:
R394.2;R730.23;R735.35
文献标志码:
A
摘要:

目的     探讨P53在纤维粘连蛋白EDA片段(fibronectin extra domain A,FN-EDA)调控结肠癌细胞凋亡中的作用及机制。方法      基因集富集分析法分析结直肠癌基因芯片EDA高表达组(EDA_high)与EDA低表达组(EDA_low)基因富集情况;干扰结肠癌SW480细胞FN-EDA表达,流式细胞术检测各组凋亡,RT-PCR检测各组凋亡相关基因RNA水平,Western blot检测各组凋亡相关蛋白表达及P53蛋白磷酸化修饰,免疫荧光检测P53亚细胞分布,蛋白质免疫共沉淀检测P53与FN-EDA相互作用。结果      结肠癌FN-EDA低表达组凋亡相关基因(KEGG_APOPTOSIS)富集上调(P<0.05);与对照组相比,FN-EDA敲低组凋亡率增加(P<0.05),P53、BAX、P21、Cleaved Caspase-3 蛋白表达上调,P53蛋白Ser15、Ser37、Ser392磷酸化水平上调,核内转位增加。FN-EDA敲低组细胞转染FN-EDA过表达慢病毒后,凋亡率降低(P<0.05),P53蛋白 Ser15、Ser37磷酸化水平下调,核内转位减少;蛋白质免疫共沉淀检测发现P53与FN-EDA具有相互作用。结论      纤维粘连蛋白EDA片段通过与P53相互作用抑制P53磷酸化和核转位,调控P53-BAX通路抑制结肠癌SW480细胞凋亡。

Abstract:

Objective      To investigate the role of fibronectin extra domain A (FN-EDA) in regulating the apoptosis of colorectal cancer cells and explore the possible mechanism. Methods     Gene set enrichment analysis (GSEA) was used to analyze the microarray data of colorectal cancer with high and low FNEDA expression. In a colorectal cancer cell line SW480 with lentiviral vector-mediated FN-EDA knockdown, the transcription levels of P53, BAX and P21 were assayed using real-time PCR, and the protein expression levels of P53, BAX, P21, Cleaved caspase-3, p-P53 (Ser15), p-P53 (Ser37) and pP53 (Ser392) were detected with Western blotting. Flow cytometry was used to detect the changes of cell apoptosis following FN-EDA knockdown, and the subcellular distribution of P53 was observed by immunofluorescence assay; the interaction between FN-EDA and P53 was analyzed using coimmunoprecipitation assay. Results     The genesets of KEGG_APOPTOSIS were obviously enriched and up-regulated in colorectal cancer cells with low FN-EDA expression (P<0.05). Compared with the control cells, the cells with FN-EDA knockdown showed significantly increased protein levels of P53, BAX, P21, Cleaved caspase-3, p-P53 (Ser15), p-P53 (Ser37) and p-P53 (Ser392) with also increased apoptotic rate (P<0.05); FN-EDA knockdown also resulted in significantly increased nuclear translocation of P53 as shown by immunofluorescence assay and Western blotting. Overexpression of FN-EDA in the cells with FN-EDA knockdown significantly inhibited the cell apoptosis (P<0.05), induced down-regulation of P53, p-P53 (Ser15), p-P53 (Ser37) and BAX, and decreased nuclear translocation of P53. Co-immunoprecipitation assay showed a direct interaction between P53 and FN-EDA. Conclusion      FN-EDA inhibits nuclear traslocation and phosphrylation of P53 via a direct interaction with P53 to regulate P53-BAX pathway and suppress the apoptosis of colorectal cancer SW480 cells.

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更新日期/Last Update: 2017-04-24