Yang Tsung-Ying, Teng Chieh-Lin Jerry, Lin Tsung-Chieh Chester, Chen Kun-Chieh, Hsu Shih-Lan, Wu Chun-Chi
Division of Chest Medicine, Department of Internal Medicine, Taichung Veterans General Hospital, Taichung, 407, Taiwan, Republic of China.
Division of Hematology/Medical Oncology, Department of Medicine, Taichung Veterans General Hospital, Taichung, Taiwan, Republic of China.
Int J Cancer. 2018 Jan 1;142(1):92-108. doi: 10.1002/ijc.31035. Epub 2017 Sep 25.
In this study, we firstly showed that p53 transcriptionally represses Aurora-A gene expression through directly binding to its promoter. DNA affinity precipitation assay and chromatin immunoprecipitation assay indicated that p53 physically bound to the Aurora-A promoter. Moreover, the in vitro and in vivo assays showed that p53 directly bound to the Aurora-A promoter together with histone deacetylase 1 (HDAC1) and mSin3a as corepressors. Furthermore, we identified that the nucleotides -360 to -354 (CCTGCCC), upstream of the Aurora-A transcriptional start site, was responsible for the p53-mediated repression. Mutation within this site disrupted its interaction with p53, mSin3a and HDAC1, as well as attenuated the repressive effect of p53 on Aurora-A promoter activity. Treatment with trichostatin A (TSA), a HDAC1 inhibitor, disrupted the interaction of p53-HDAC1-mSin3a complex with the nucleotides -365∼-345 region, and enhanced the Aurora-A promoter activity and gene expression. Additionally, knockdown of p53 or mSin3a also drastically blocked the formation of p53-HDAC1-mSin3a repressive complex onto this promoter region and elevated the Aurora-A promoter activity and gene expression. Moreover, the p53-HDAC1-mSin3a repressive complex also involved in the inhibition of Aurora-A gene expression upon cisplatin treatment. Finally, the clinical investigation showed that Aurora-A and p53 exhibited an inverse correlation in both the expression level and prognostic status, and the low p53/high Aurora-A showed the poorest prognosis of NSCLC patients. Our findings showed novel regulatory mechanisms of p53 in regulating Aurora-A gene expression in NSCLC cells.
在本研究中，我们首先表明p53通过直接结合Aurora-A基因启动子转录抑制其基因表达。DNA亲和沉淀试验和染色质免疫沉淀试验表明p53与Aurora-A启动子发生物理结合。此外，体外和体内试验表明，p53与组蛋白去乙酰化酶1（HDAC1）和mSin3a作为共抑制因子直接结合到Aurora-A启动子上。此外，我们确定Aurora-A转录起始位点上游的核苷酸-360至-354（CCTGCCC）负责p53介导的抑制作用。该位点的突变破坏了其与p53、mSin3a和HDAC1的相互作用，以及减弱了p53对Aurora-A启动子活性的抑制作用。用HDAC1抑制剂曲古抑菌素A（TSA）处理破坏了p53-HDAC1-mSin3a复合物与核苷酸-365∼-345区域的相互作用，并增强了Aurora-A启动子活性和基因表达。此外，敲低p53或mSin3a也显著阻断了p53-HDAC1-mSin3a抑制复合物在该启动子区域的形成，并提高了Aurora-A启动子活性和基因表达。此外，p53-HDAC1-mSin3a抑制复合物也参与顺铂处理后对Aurora-A基因表达的抑制。最后，临床研究表明，Aurora-A和p53在表达水平和预后状态上均呈负相关，低p53/高Aurora-A的非小细胞肺癌患者预后最差。我们的研究结果揭示了p53在调节非小细胞肺癌细胞中Aurora-A基因表达方面的新调控机制。
