Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, Hangzhou 310058, PR China.
Environ Toxicol Pharmacol. 2010 Jan;29(1):79-86. doi: 10.1016/j.etap.2009.11.001. Epub 2009 Nov 5.
Human Pol ι is a highly distributed, low-fidelity DNA polymerase lacking intrinsic exonuclease proofreading activity, thus its effects are strictly regulated. We predicted and cloned the promoter region of the human POLI gene. Successively, by transfection of deletion constructs of the POLI promoter, we demonstrated that the regions -848/-408 and -30/+215 contained positive regulatory elements, and the region +215/+335 had proximal promoter activity. Overexpression of Sp1 significantly increased the transcriptional activity of the promoter, and mutation of the Sp1 site reversed Sp1-induced promoter transactivation. Quantitative RT-PCR showed that POLI mRNA expression was up-regulated in human amnion FL cells treated by the carcinogen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Reporter gene assays demonstrated that MNNG also significantly increased the transcriptional activity of the predicted promoter (-848/+335) and the proximal promoter (+215/+335). However, the promoter with the Sp1 site mutation had no response to MNNG treatment, suggesting that Sp1 plays an important role in the transcriptional regulation of the POLI gene stimulated by MNNG. Our data suggest that abnormal regulation of Pol ι may be involved in the mutagenesis and carcinogenesis induced by environmental chemicals.
人源 Pol ι 是一种分布广泛但保真度低的 DNA 聚合酶，缺乏内在的核酸外切酶校对活性，因此其功能受到严格调控。我们预测并克隆了人 POLI 基因的启动子区域。通过转染 POLI 启动子缺失构建体，我们证明了 -848/-408 和 -30/+215 区域包含正调控元件，+215/+335 区域具有近端启动子活性。Sp1 的过表达显著增加了启动子的转录活性，而 Sp1 位点的突变则逆转了 Sp1 诱导的启动子反式激活。定量 RT-PCR 显示，致癌剂 N-甲基-N'-硝基-N-亚硝基胍（MNNG）处理人羊膜 FL 细胞后，POLI mRNA 表达上调。报告基因分析表明，MNNG 还显著增加了预测启动子（-848/+335）和近端启动子（+215/+335）的转录活性。然而，Sp1 位点突变的启动子对 MNNG 处理没有反应，这表明 Sp1 在 MNNG 刺激的 POLI 基因转录调控中发挥重要作用。我们的数据表明，Pol ι 的异常调节可能参与了环境化学物质诱导的突变和致癌作用。
