G-quadruplex is a DNA secondary structure that has been shown to play an important role in biological systems. In a previous study, we identified 1998 G-quadruplex-forming sequences using a mouse CpG islands DNA microarray with a fluorescent-labeled G-quadruplex ligand. Among these putative G-quadruplex-forming sequences, G-quadruplex formation was verified for 10 randomly selected sequences by CD spectroscopy and DMS footprinting analysis. In this study, the biological function of the 10 G-quadruplex-forming sequences in the transcriptional regulation has been analyzed using a reporter assay. When G-quadruplex-forming sequences from the Dele and Cdc6 genes have been cloned in reporter vectors carrying a minimal promoter and the luciferase gene, luciferase expression is activated. This has also been detected in experiments applying a promoterless reporter vector. Mutational analysis reveals that guanine bases, which form the G-tetrads, are important in the activation. In addition, the activation has been found to decrease by the telomestatin derivative L1H1-7OTD which can bind to the G-quadruplex DNA. When Dele and Cdc6 CpG islands, containing the G-quadruplex-forming sequence, have been cloned in the promoterless reporter vector, the luciferase expression is activated. Mutational analysis reveals that the expression level is decreased by mutation on Dele G-quadruplex; however, increased by mutation on Cdc6 G-quadruplex. Dele and Cdc6 G-quadruplex formation is significant in the transcriptional regulation. Dele and Cdc6 G-quadruplex DNA alone possess enhancer and promotor function. When studied in more complex CpG islands Dele G-quadruplex also demonstrates promotor activity, whereas Cdc6 G-quadruplex may possess a dual function of transcriptional regulation.

中文翻译：

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鉴定在Dele和Cdc6 CpG岛中具有转录调节功能的G-四链体结构。

G-四链体是DNA二级结构，已显示在生物系统中起重要作用。在先前的研究中，我们使用带有荧光标记的G-四链体配体的小鼠CpG岛DNA微阵列鉴定了1998年G-四链体形成序列。在这些推定的G-四链体形成序列中，通过CD光谱和DMS足迹分析对10个随机选择的序列验证了G-四链体形成。在这项研究中，已使用报告基因分析法分析了10个G-四链体形成序列在转录调控中的生物学功能。当已经将来自Dele和Cdc6基因的G-四链体形成序列克隆到携带最小启动子和荧光素酶基因的报告载体中时，荧光素酶表达被激活。在使用无启动子报告载体的实验中也已检测到这一点。突变分析表明，形成G-四联体的鸟嘌呤碱基在激活中很重要。另外，已经发现可以与G-四链体DNA结合的端粒他汀衍生物L1H1-7OTD降低了活化。当包含G-四链体形成序列的Dele和Cdc6 CpG岛已克隆到无启动子的报告载体中时，萤光素酶表达被激活。突变分析表明，在Dele G-quadruplex上，表达水平降低。但是，由于Cdc6 G四联体的突变而增加。Dele和Cdc6 G四联体的形成在转录调控中很重要。Dele和Cdc6 G-四链体DNA单独具有增强子和启动子功能。