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

转录因子ATF5的新功能：阻断辐射诱导的P53依赖的细胞凋亡。目的：P53依赖的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，携带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的肿瘤细胞也对电离辐射表现出不同程度的辐射敏感性。这表明还有其他途径影响依赖于P53的细胞死亡机制。在这里，我们描述了一种新的分子，它通过抑制p53的反式激活活性来抑制辐射诱导的细胞凋亡。材料和方法：我们用10Gy射线照射小鼠可移植的恶性细胞株QRsP细胞，并从存活的克隆中建立了24个亚克隆。这些克隆细胞被再次照射，最敏感的克隆QRsPIR-5被用于本实验。所有子…更多的克隆细胞具有与亲本QRsP细胞相同的形态特征，且各克隆之间均未发现P53突变。克隆形成实验表明，QRsPIR-5细胞在10Gy剂量下的存活分数不到亲代QRsP细胞的20%。流式细胞仪分析还显示，携带野生型p53的重组腺病毒(Ad-P53)感染QRsPIR-5细胞后，细胞的凋亡率高于亲本细胞(26.5%vs.7.1%)。有趣的是，亲本细胞和QRsPIR-5细胞在移植实验中具有相同程度的致瘤性。结果：综合cDNA阵列分析表明，亲本细胞和QRsPIR-5细胞在体外和体内的基因表达存在差异。其中，23个基因在亲本细胞和QRsP-5细胞的体外和体内差异表达具有较高的严格阈值(小于0.5或大于2.0)。一个这样的基因，bZIP转录因子ATF5，可能解释了辐射敏感性的差异。外源ATF5的表达使QRsPIR-5细胞具有与亲本细胞相似的辐射抗性水平(集落试验)。此外，QRsPIR-5细胞对Ad-P53诱导的细胞凋亡具有抵抗力。荧光素酶报告实验表明，过表达的ATF5显著抑制了野生型p53的转录活性。有趣的是，时间推移分析表明，ATF5转基因的QRsPIR-5细胞的运动速度加快。结论：ATF5可能是一种有效的p53抑制因子。ATF5在肿瘤中的高表达可能与恶性表型的增强有关，如放射抗性或更大的细胞运动性。较少

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