[Phosphorylation status of ASPP2 modulates p53 apoptotic function in oxaliplatin-induced apoptosis of colorectal cancer HCT116 cells].
[Phosphorylation status of ASPP2 modulates p53 apoptotic function in oxaliplatin-induced apoptosis of colorectal cancer HCT116 cells].
复制标题
DOI：
10.3760/cma.j.issn.0253-3766.2014.06.004
复制
发表时间：
2014-06
期刊：
影响因子：
--
通讯作者：
Q. Hou;Hongwei Zhao;Weipeng Gong;Zhenyu Zhu;Yue Han;Dexi Chen;Hongliang Guo
中科院分区：
文献类型：
--
作者：
Q. Hou;Hongwei Zhao;Weipeng Gong;Zhenyu Zhu;Yue Han;Dexi Chen;Hongliang Guo
目的探讨p53凋亡刺激蛋白2（ASPP 2）磷酸化状态在ASPP 2-p53凋亡通路活性调节中的作用。方法分别将编码绿色荧光蛋白（GFP）的质粒、编码组成型非磷酸化ASPP 2（Am）的质粒和编码野生型ASPP 2（Aw）的质粒转染细胞，使其分别过表达磷酸化和非磷酸化ASPP 2蛋白。奥沙利铂诱导细胞凋亡。用FITC标记的Annexin V和PI染色后，用流式细胞仪测定细胞凋亡率。Western blot检测ASPP 2蛋白表达水平及磷酸化状态。免疫沉淀法观察ASPP 2与p53的相互作用。结果奥沙利铂可诱导HCT 116细胞凋亡，并引起ASPP 2的ser 92/ser 361磷酸化。转染Aw和Am质粒的细胞凋亡率分别为（3.8 ± 1.0）%和（3.9 ± 1.2）%，与转染GFP质粒的细胞凋亡率[（4.0 ± 0.8）%]相比，差异无统计学意义（P > 0.05）。奥沙利铂处理后，Aw质粒转染的细胞凋亡率为（46.7 ± 3.9）%，显著高于Am和GFP质粒转染的细胞[Am和GFP质粒转染的细胞凋亡率分别为（40.1 ± 10.2）%和（37.1 ± 6.9）%，P0.05]。这些结果表明，磷酸化ASPP 2通过p53依赖性途径促进奥沙利铂诱导的HCT 116细胞凋亡。ASPP 2的磷酸化状态影响其与p53的结合活性。结论ASPP 2磷酸化状态在奥沙利铂诱导大肠癌HCT 116细胞凋亡中调节p53凋亡功能。
OBJECTIVE To investigate the role of apoptosis stimulating protein 2 of p53 (ASPP2) phosphorylation status in the regulation of ASPP2-p53 apoptotic pathway activity. METHODS Cells were individually transfected with green fluorescent protein (GFP)-encoding vector, constitutively non-phosphorylatable ASPP2 mutant-ASPP2 (Am)-encoding vector, and wild type ASPP2 (Aw)-encoding vector) plasmids, respectively, to make them overexpressing phosphorylated and non-phosphorylated ASPP2 proteins, respectively. Cell apoptosis was induced by oxaliplatin. The apoptosis rate of cells was determined by flow cytometry after staining with FITC-conjugated annexin V and PI. ASPP2 protein level and its phosphorylation status were observed by Western blot. The interaction between ASPP2 and p53 was observed by immunoprecipitation assay. RESULTS Oxaliplatin induced cell apoptosis and caused phosphorylation of ASPP2 at ser92/ser361 in the HCT116 cells. The apoptosis rate of Aw and Am plasmids-transfected cells were (3.8 ± 1.0)% and (3.9 ± 1.2)% respectively, statistically with a non-significant difference (P > 0.05) in comparison with that of the GFP plasmid-transfected cells [(4.0 ± 0.8)%]. After oxaliplatin treatment, the apoptosis rate of Aw plasmid-transfected cells was (46.7 ± 3.9)%, significantly higher than that of the Am and GFP plasmid-transfected cells [(40.1 ± 10.2)% and (37.1 ± 6.9)%, respectively, P 0.05) between Am and GFP plasmid-transfected cells. These results indicate that phosphorylated ASPP2 promoted the oxaliplatin-induced apoptosis of HCT116 cells through a p53-dependent pathway. Phosphorylation status of ASPP2 influenced its binding activity to p53. CONCLUSION Phosphorylation status of ASPP2 modulates p53 apoptotic function in oxaliplatin-induced apoptosis of colorectal cancer HCT116 cells.