Contrasting genome-wide distribution of 8-hydroxyguanine and acrolein-modified adenine during oxidative stress-induced renal carcinogenesis
Contrasting genome-wide distribution of 8-hydroxyguanine and acrolein-modified adenine during oxidative stress-induced renal carcinogenesis
复制标题
DOI：
10.2353/ajpath.2006.051280
复制
发表时间：
2006-10-01
影响因子：
6
通讯作者：
Toyokuni, Shinya
中科院分区：
文献类型：
--
作者：
Akatsuka, Shinya;Aung, Than Tin;Toyokuni, Shinya
氧化应激是对基因组的持续威胁，并与人类死亡的主要原因有关，包括癌症、动脉粥样硬化和衰老。在这里，我们建立了一种方法，通过使用特异性单克隆抗体免疫沉淀酶消化的基因组 DNA，生成含有氧化修饰碱基的基因组 DNA 片段文库。我们在次氮基三乙酸铁诱导的小鼠肾癌模型中将该技术应用于两种不同的碱基修饰：8-羟基鸟嘌呤和 1,N-6-丙腺嘌呤（阿克罗蛋白-Ade）。对来自 10 至 12 周龄雄性 C57BL/6 小鼠、未经治疗的对照小鼠或腹腔注射 3 mg 铁/kg 次氮基三乙酸铁 6 小时后的肾皮质基因组 DNA 进行酶消化、免疫沉淀、克隆并定位到每条染色体。结果显示，两个修饰碱基的分布不是随机的，而是在染色体、基因大小和表达方面存在差异，这可以部分地用染色体区域来解释。在野生型小鼠中，GC含量低的区域更有可能含有这两种修饰碱基。通过定量聚合酶链式反应分析确定，敲除 OGG1（一种基因组 8-羟基鸟嘌呤的修复酶）会增加丙烯醛-Ade 的含量。这种多功能技术将引入一个新的研究领域，作为氧化应激下关键基因组位点的高通量筛选方法。
Oxidative stress is a persistent threat to the genome and is associated with major causes of human mortality, including cancer, atherosclerosis, and aging. Here we established a method to generate libraries of genomic DNA fragments containing oxidatively modified bases by using specific monoclonal antibodies to iinmunoprecipitate enzyme-digested genome DNA. We applied this technique to two different base modifications, 8-hydroxyguanine and 1,N-6-propanoadenine (acrotein-Ade), in a ferric nitrllotriacetate-induced murine renal carcinogenesis model. Renal cortical genomic DNA derived from 10- to 12-week-old male C57BL/6 mice, of untreated control or 6 hours after intraperitoneal injection of 3 mg iron/kg ferric nitrilotriacetate, was enzyme digested, immunoprecipitated, cloned, and mapped to each chromosome. The results revealed that distribution of the two modified bases was not random but differed in terms of chromosomes, gene size, and expression, which could be partially explained by chromosomal territory. In the wild-type mice, low GC content areas were more likely to harbor the two modified bases. Knockout of OGG1, a repair enzyme for genomic 8-hydroxyguanine, increased the amounts of acrolein-Ade as determined by quantitative polymerase chain reaction analyses. This versatile technique would introduce a novel research area as a high-throughput screening method for critical genomic loci under oxidative stress.