Park Chang Won, Kren Betsy T, Largaespada David A, Steer Clifford J
Department of Medicine, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA.
Genes Cells. 2005 Aug;10(8):763-76. doi: 10.1111/j.1365-2443.2005.00875.x.
The Sleeping Beauty transposon is a recently developed non-viral vector that can mediate insertion of transgenes into the mammalian genome. Foreign DNA elements that are introduced tend to invoke a host-defense mechanism resulting in epigenetic changes, such as DNA methylation, which may induce transcriptional inactivation of mammalian genes. To assess potential epigenetic modifications associated with Sleeping Beauty transposition, we investigated the DNA methylation pattern of transgenes inserted into the mouse genome as well as genomic regions flanking the insertion sites with bisulfite-mediated genomic sequencing. Transgenic mouse lines were created with two different Sleeping Beauty transposons carrying either the Agouti or eGFP transgene. Our results showed that DNA methylation in the keratin-14 promoter and Agouti transgene were negligible. In addition, two different genomic loci flanking the Agouti insertion site exhibited patterns of DNA methylation similar to wild-type mice. In contrast, high levels of DNA methylation were observed in the eGFP transgene and its ROSA26 promoter. These results indicate that transposition via Sleeping Beauty into the mouse genome may result in a significant level of de novo DNA methylation. This may depend on a number of different factors including the cargo DNA sequence, chromosomal context of the insertion site, and/or host genetic background.
睡美人转座子是一种最近开发的非病毒载体，它能够介导转基因插入哺乳动物基因组。引入的外源DNA元件往往会引发宿主防御机制，导致表观遗传变化，如DNA甲基化，这可能会诱导哺乳动物基因的转录失活。为了评估与睡美人转座相关的潜在表观遗传修饰，我们用亚硫酸氢盐介导的基因组测序研究了插入小鼠基因组的转基因以及插入位点侧翼基因组区域的DNA甲基化模式。用携带刺鼠基因或增强绿色荧光蛋白（eGFP）转基因的两种不同睡美人转座子创建了转基因小鼠品系。我们的结果表明，角蛋白-14启动子和刺鼠基因中的DNA甲基化可以忽略不计。此外，刺鼠基因插入位点侧翼的两个不同基因组位点表现出与野生型小鼠相似的DNA甲基化模式。相比之下，在eGFP转基因及其ROSA26启动子中观察到高水平的DNA甲基化。这些结果表明，通过睡美人转座进入小鼠基因组可能会导致显著水平的从头DNA甲基化。这可能取决于许多不同因素，包括货物DNA序列、插入位点的染色体背景和/或宿主遗传背景。
