Nm23-H1 Protein Binds to APE1 at AP Sites and Stimulates AP Endonuclease Activity Following Ionizing Radiation of the Human Lung Cancer A549 Cells
Nm23-H1 Protein Binds to APE1 at AP Sites and Stimulates AP Endonuclease Activity Following Ionizing Radiation of the Human Lung Cancer A549 Cells
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Nm23-H1 蛋白在人肺癌 A549 细胞电离辐射后与 AP 位点的 APE1 结合并刺激 AP 核酸内切酶活性
DOI：
10.1007/s12013-011-9238-9
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发表时间：
2011-12-01
影响因子：
2.6
通讯作者：
Xiang, De-Bing
中科院分区：
文献类型：
--
作者：
Zhang, Zhi-Min;Yang, Xue-Qin;Xiang, De-Bing
非转移性蛋白 23 同源物 1 (Nm23-H1) 是一种具有 DNase 和组氨酸蛋白激酶活性的多功能蛋白。人无嘌呤核酸内切酶-1 (APE1) 是 AP 核酸内切酶 DNA 碱基切除修复 (BER) 酶，参与多种重要的细胞功能。由于Nm23-H1和APE1蛋白之间的关系尚不清楚，我们在用X射线照射人肺癌A549细胞后的不同时间点评估了它们的相互作用。我们发现 Nm23-H1 和 APE1 过度表达是由辐射以剂量和时间依赖性方式诱导的。辐射后两种蛋白质的亚细胞分布模式发生逆转。照射后，A549细胞中最初表现为核定位的APE1在细胞质中逐渐增多，而主要表现为细胞质定位的Nm23-H1在细胞核中逐渐增多。 Nm23-H1 和 APE1 相互作用通过 His-pull-down 和免疫共沉淀测定得到证实。通过对含有 AP 位点的 DNA 片段进行 DNA 亲和沉淀分析，也检测到 X 射线照射的 A549 细胞中是否存在 Nm23-H1/APE1 复合物。虽然Nm23-H1的AP内切酶活性太弱而无法检测到，但APE1的AP内切酶活性随着Nm23-H1表达的增强而增加。总之，我们的数据指出了 Nm23-H1 通过 DNA BER 酶 APE1 的参与来保护细胞免受氧化应激的机制。
Non-metastatic protein-23 homolog-1 (Nm23-H1) is a multifunctional protein with DNase and histidine protein kinase activities. Human apurinic endonuclease-1 (APE1) is the AP endonuclease DNA base excision repair (BER) enzyme involved in several important cellular functions. Since the relationship between Nm23-H1 and APE1 proteins is unclear, we evaluated their interaction at different time points after irradiating human lung cancer A549 cells with X-rays. We found that Nm23-H1 and APE1 overexpression was induced by irradiation in a dose- and time-dependent manner. Subcellular distribution pattern of both proteins was reversed after irradiation. After irradiation, APE1 that initially showed nuclear localization was gradually increased in the cytoplasm, whereas Nm23-H1 that mainly showed cytoplasmic localization was gradually increased in the nuclei of A549 cells. Nm23-H1 and APE1 interaction was demonstrated by His-pull-down and co-immunoprecipitation assays. The presence of Nm23-H1/APE1 complex in X-ray-irradiated A549 cells was also detected by DNA affinity precipitation analysis of a DNA fragment containing an AP site. Although the AP endonuclease activity of Nm23-H1 was too weak to be detected, the AP endonuclease activity of APE1 was increased with the enhanced Nm23-H1 expression. In conclusion, our data point to a mechanism by which Nm23-H1 protects cells against oxidative stress through the engagement of DNA BER enzyme APE1.