Liu Xiangguo, Yue Ping, Khuri Fadlo R, Sun Shi-Yong
Department of Hematology and Oncology, Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA 30322, USA.
Cancer Res. 2005 Oct 15;65(20):9169-75. doi: 10.1158/0008-5472.CAN-05-0939.
Decoy receptor 2 (DcR2) is one of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors and suppresses TRAIL-induced apoptosis. Its expression, like the other three TRAIL receptors (i.e., DR4, DR5, and DcR1), is regulated by p53. Here, we report that DcR2 is a p53 target gene and regulates chemosensitivity. In this study, we identified a p53-binding site (p53BS) in the first intron of the DcR2 gene. This p53BS is almost identical to the ones found in the first introns of other three TRAIL receptor genes. By a chromatin immunoprecipitation assay, we detected that the p53 protein bound to the DcR2 p53BS in intact cells. Subcloning of the DcR2 p53BS into a luciferase reporter vector driven by a SV40 promoter exhibited enhanced luciferase activity when transiently cotransfected with a wild-type (wt) p53 expression vector in p53-null cell lines or stimulated with DNA-damaging agents in cell lines having wt p53. Moreover, when the DcR2 p53BS, together with its own corresponding promoter regions, was subcloned into a basic luciferase vector without a promoter element, its transcriptional activities were strikingly increased by cotransfection of the wt p53 gene. However, when this p53BS was deleted from the construct, wt p53 failed to transactivate this reporter construct. Collectively, we conclude that p53 directly regulates the DcR2 gene expression via an intronic p53BS. In addition, overexpression of DcR2 conferred resistance to TRAIL-mediated apoptosis and attenuated cell response to DNA-damaging agents, whereas silencing of DcR2 expression enhanced chemotherapeutic agent-induced apoptosis. These results suggest that DcR2 regulates chemosensitivity.
诱饵受体2（DcR2）是肿瘤坏死因子相关凋亡诱导配体（TRAIL）受体之一，可抑制TRAIL诱导的细胞凋亡。其表达与其他三种TRAIL受体（即DR4、DR5和DcR1）一样，受p53调控。在此，我们报告DcR2是一个p53靶基因并调节化学敏感性。在本研究中，我们在DcR2基因的第一个内含子中鉴定出一个p53结合位点（p53BS）。这个p53BS与在其他三个TRAIL受体基因的第一个内含子中发现的位点几乎相同。通过染色质免疫沉淀试验，我们检测到在完整细胞中p53蛋白与DcR2的p53BS结合。将DcR2的p53BS亚克隆到由SV40启动子驱动的荧光素酶报告载体中，当在p53缺失的细胞系中与野生型（wt）p53表达载体共转染或在具有wt p53的细胞系中用DNA损伤剂刺激时，荧光素酶活性增强。此外，当将DcR2的p53BS及其自身相应的启动子区域亚克隆到一个没有启动子元件的基本荧光素酶载体中时，通过共转染wt p53基因，其转录活性显著增加。然而，当从构建体中删除这个p53BS时，wt p53无法激活这个报告构建体。总体而言，我们得出结论，p53通过一个内含子p53BS直接调节DcR2基因表达。此外，DcR2的过表达赋予对TRAIL介导的细胞凋亡的抗性，并减弱细胞对DNA损伤剂的反应，而DcR2表达的沉默增强化疗药物诱导的细胞凋亡。这些结果表明DcR2调节化学敏感性。
