天津医药

天津医药 ›› 2019, Vol. 47 ›› Issue (1): 5-9.doi: 10.11958/20181312

• 细胞与分子生物学 • 上一篇    下一篇

Aurora-B介导NPM1蛋白磷酸化促进骨肉瘤细胞 恶性表型的体外研究

皮闻森，刘家明，黄山虎，刘志礼△   

  1. 基金项目:国家自然科学基金资助项目（81560435）；江西省杰出青年人才资助项目（20162BCB23057）；江西省自然科学基金资助项目 （20161ACB20011） 作者单位：南昌大学第一附属医院骨科（邮编330006） 作者简介：皮闻森（1992），男，硕士在读，主要从事骨肉瘤肺转移分子机制的研究 △通讯作者 E-mail: zgm7977@163.com
  • 收稿日期:2018-08-31 修回日期:2018-11-15 出版日期:2019-01-15 发布日期:2019-01-15
  • 通讯作者: 刘志礼 E-mail:zgm7977@163.com
  • 基金资助:
    国家自然科学基金;江西省杰出青年人才资助项目;江西省自然科学基金资助项目;南昌大学研究生创新专项资金项目

Aurora-B-mediated phosphorylation of NPM1 promotes malignant phenotype of OS cells in vitro

PI Wen-sen, LIU Jia-ming, HUANG Shan-hu, LIU Zhi-li△   

  1. Department of Orthopedics, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China △Corresponding Author E-mail: zgm7977@163.com
  • Received:2018-08-31 Revised:2018-11-15 Published:2019-01-15 Online:2019-01-15
  • Contact: Zhi-Li LIU E-mail:zgm7977@163.com

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摘要： 目的 探讨 Aurora-B 介导 NPM1（nucleophosmin1）蛋白磷酸化水平改变对骨肉瘤细胞恶性表型的影响。 方法 运用生物信息学数据库预测NPM1在肉瘤中的表达情况以及Aurora-B与NPM1的相互关系。构建重组慢病 毒载体，获得沉默 Aurora-B 慢病毒（LV/ShAurora-B）、过表达 NPM1 慢病毒（LV/NPM1）和阴性对照慢病毒（LV/ negative)。转染人源骨肉瘤细胞系143B及U2-OS细胞，分为Lv/ShAurora-B组、NC（LV/negative）组和LV/ShAuroraB+LV/NPM1共转染组。Western blot分别检测LV/ShAurora-B组和NC组中Aurora-B、磷酸化NPM1ser125蛋白表达。采 用CCK-8法检测3组细胞增殖情况，Wound healing 法检测细胞迁移能力，Transwell invasion 法检测细胞侵袭能力。 结果 生物信息学结果显示，NPM1在肉瘤中高表达以及NPM1高表达患者的预后较差，且NPM1存在丝氨酸和苏氨 酸磷酸化位点，Aurora-B与NPM1之间可能存在磷酸化作用。Western blot结果显示，与NC组相比，LV/ShAurora-B组 中Aurora-B和磷酸化NPM1ser125蛋白的表达均降低（P＜0.05），而NPM1蛋白差异无统计学意义（P＞0.05）。体外表型 实验显示，LV/ShAurora-B组细胞的增殖、迁徙和侵袭能力均低于NC组（P＜0.05），且LV/ShAurora-B+LV/NPM1共转 染组能部分恢复下调Aurora-B对骨肉瘤细胞恶性表型的抑制（P＜0.05）。结论 Aurora-B通过介导NPM1蛋白磷酸 化促进骨肉瘤细胞恶性表型。

关键词: 骨肉瘤, 计算生物学, Aurora-B, NPM1, 磷酸化NPM1ser125, 恶性表型

Abstract: Objective To investigate the effect of Aurora-B-mediated phosphorylation of nucleophosmin1 (NPM1) protein on the malignant phenotype of osteosarcoma (OS) cells. Methods Bioinformatic prediction was performed to investigate the expression level of NPM1 in sarcoma, and the potential relationship between Aurora-B and NPM1. OS cells (U2-OS and 143B）were transfected with LV/ShAurora-B, NC-LV/ShAurora-B, and co-infected by LV/ShAurora-B+NC LV/NPM1. Transwell invasion, Wound healing and CCK8 assays were performed to evaluate the malignant phenotype of OS cells. Results Bioinformatic prediction showed that NPM1 protein was overexpression in sarcoma tissues, and Aurora-B may mediate phosphorylation of NPM1. The Western blot results revealed that the expression levels of Aurora-B and pNPM1ser125 protein were significantly lower in cells infected with LV/ShAurora-B than those in cells treated with Lv/negative (P＜0.05). Interesting, the expression levels of NPM1 protein in cells infected by LV/ShAurora-B were similar to those in cells infected by NC-LV/ShAurora-B. The invaded cells, migration rate and the proliferation of OS cells were significantly reduced in cells down-regulated Aurora-B than those in negative group (P＜0.05). However, the inhibiting effect was partially restored by up-regulation of NPM1 in OS cells. Conclusion Aurora-B-mediated phosphorylation of NPM1 promotes malignant phenotype of OS cells in vitro.

Key words: osteosarcoma, computational biology, Aurora-B, NPM1, pNPM1ser125, malignant phenotype