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Radiation-induced highly-malignant cancer cells: the mechanism and how to deal with it

辐射诱发高度恶性癌细胞：机制及处理方法

基本信息

批准号：
18390325
负责人：
NISHIOKA Takeshi
金额：
$ 6.24万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18390325/
关键词：
p53 apoptosis irradiation radiosensitivity transcription factor ATF5

项目摘要

Novel function of transcription factor ATF5: blockade of p53-dependent apoptosis induced by irradiation.Purpose: p53-dependent cell death is considered as a predominant mechanism of tumor cell apoptosis induced by ionizing irradiation, and a large number of studies have shown that mutant p53-harboring tumor cells with a p53 gene mutation exhibit radioresistance. However, even tumor cells that express wild-type p53 display various degrees of radiosensitivity to ionizing irradiation. This indicates that there are additional pathways that affect p53-dependent cell death mechanisms. Here we describe a novel molecule that represses radiation-induced apoptosis by inhibiting trans-activation activity of p53.Materials and Methods: We irradiated QRsP cells, a mouse transplantable malignant cell line, at 10Gy and from surviving colonies 24 sub-clones were established. These clonal cells were re-irradiated and the most readio-sensitive clone, QRsPIR-5, was used in the current experiment. All sub- … More cloned cells had the same morphological appearances as the parental QRsP cells and there were no p53 mutations of among any of the clones. Colony assay indicated that the survival fraction of QRsPIR-5 cells at a dose of 10 Gy was less than 20% of the survival fraction of the parental QRsP cells. Flow cytometer analysis also indicated a higher apoptotic index after infection with recombinant adenovirus containing wild-type p53 (Ad-p53) in QRsPIR-5 cells compared with the parent cell (26.5% vs. 7.1%). Interestingly, the parental and QRsPIR-5 cells had the same degree of tumorigenicity in a transplant experiment.Results: Comprehensive cDNA array analyses demonstrated differential gene expressions between the parental and QRsPIR-5 cells, both in vitro and in vivo. Among these, 23 genes were expressed differently both in vitro and in vivo between the parent and QRsP-5 cells with a high stringent threshold (less than 0.5 or more than 2.0). One such gene, bZIP transcription factor ATF5, which might explain difference in radio-sensitivity. Exogenous expression of ATF5 gave QRsPIR-5 cells a radioresistance level similar to that of the parental cells (colony assay). Moreover, QRsPIR-5 cells gained resistance to Ad-p53-induced apoptosis. A luciferase reporter assay demonstrated that over-expressed ATF5 repressed the transcriptional activity of wild-type p53 drastically. Interestingly, time lapse analysis indicated accelerated motility in ATF5-transfected QRsPIR-5 cells.Conclusion: It is likely that ATF5 is a potent repressor of p53. Elevated expression of ATF5 in a tumor may relate to enhanced malignant phenotypes, such as radio-resistance or greater cell motility. Less

目的：p53依赖性细胞死亡被认为是电离辐射诱导肿瘤细胞凋亡的主要机制，大量研究表明，p53基因突变的肿瘤细胞具有辐射抗性。然而，即使是表达野生型p53的肿瘤细胞也显示出对电离辐射的不同程度的辐射敏感性。这表明有其他途径影响p53依赖性细胞死亡机制。材料和方法：用10 Gy的γ射线照射小鼠可移植性恶性肿瘤细胞系QRsP细胞，从存活的集落中建立24个亚克隆。将这些克隆细胞再照射，并将最放射敏感的克隆QRsPIR-5用于当前实验。所有子- ...更多信息克隆的细胞具有与亲本QRsP细胞相同的形态学外观，并且在任何克隆中没有p53突变。集落形成实验表明，QRsPIR-5细胞在10戈伊剂量下的存活率低于亲本QRsP细胞的20%。流式细胞仪分析还表明，与亲本细胞相比，用含有野生型p53的重组腺病毒（Ad-p53）感染QRsPIR-5细胞后，QRsPIR-5细胞中的凋亡指数更高（26.5%对7.1%）。有趣的是，父母和QRsPIR-5细胞有相同程度的致瘤性在移植experiments.Results：全面的cDNA阵列分析表明，父母和QRsPIR-5细胞之间的差异基因表达，在体外和体内。其中，23个基因在亲本和QRsP-5细胞之间在体外和体内都有不同的表达，具有高严格阈值（小于0.5或大于2.0）。一个这样的基因，bZIP转录因子ATF 5，这可能解释了辐射敏感性的差异。ATF 5的外源性表达给予QRsPIR-5细胞与亲本细胞相似的辐射抗性水平（集落测定）。此外，QRsPIR-5细胞对Ad-p53诱导的凋亡具有抵抗性。荧光素酶报告基因检测表明，过度表达的ATF 5抑制野生型p53的转录活性急剧。有趣的是，时间推移分析表明，加速运动在ATF 5转染QRsPIR-5 cells.Conclusion：这是可能的，ATF 5是一个有效的阻遏p53。肿瘤中ATF 5的表达升高可能与恶性表型增强有关，例如放射抗性或更大的细胞运动性。少