CpG content affects gene silencing in mice: evidence from novel transgenes
CpG content affects gene silencing in mice: evidence from novel transgenes
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DOI：
10.1186/gb-2003-4-9-r53
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发表时间：
2003-01-01
期刊：
影响因子：
12.3
通讯作者：
Nicolas, JF
中科院分区：
文献类型：
--
作者：
Chevalier-Mariette, C;Henry, I;Nicolas, JF
背景：转基因通常是利用远亲基因组中的调控元件来设计的。尽管即使在转基因小鼠中也经常能获得正确的表达模式，但不适当的表达，特别是在广泛表达的基因启动子的情况下，已经有报道。结果：我们描述了一种新的分析表观遗传控制的方法，该方法通过将CpG含量从高到零的LacZ报告基因与一个广泛表达的基因-翻译延长因子I的α亚基的CpG岛启动子相结合，直接检测CPGS的参与。我们的数据表明，CpG含量为零的LacZ转基因基因取消了在CpG含量较高的转基因品系的体细胞组织中观察到的强烈的转基因抑制。对转基因表达和甲基化模式的研究表明，在基因组的从头甲基化过程中，含有CpG岛的启动子只有在与CpG零基因结合时才能逃脱甲基化。在其他转基因株系中，启动子的甲基化可能导致了转录沉默。结论：我们证明了转基因转录区域中CpG序列的密度可能在转录抑制中起到因果作用。这些结果表明，CpG岛逃避从头甲基化的机制对相邻序列的CpG密度很敏感。这些发现对于设计用于控制表达的转基因具有重要意义。
Background: Transgenes are often engineered using regulatory elements from distantly related genomes. Although correct expression patterns are frequently achieved even in transgenic mice, inappropriate expression, especially with promoters of widely expressed genes, has been reported. DNA methylation has been implicated in the aberrant expression, but the mechanism by which the methylation of a CpG-rich sequence can perturb the functioning of a promoter is unknown.Results: We describe a novel method for analyzing epigenetic controls that allows direct testing of CpGs involvement by using LacZ reporter genes with a CpG content varying from high to zero that are combined with a CpG island-containing promoter of a widely expressed gene - the alpha-subunit of the translation elongation factor I. Our data revealed that a LacZ transgene with null CpG content abolished the strong transgene repression observed in the somatic tissues of transgenic lines with higher CpG content. Investigation of transgene expression and methylation patterns suggests that during de novo methylation of the genome the CpG island-containing promoter escapes methylation only when combined with the CpG-null transgene. In the other transgenic lines, methylation of the promoter may have led to transcriptional silencing.Conclusions: We demonstrate that the density of CpG sequences in the transcribed regions of transgenes can have a causal role in repression of transcription. These results show that the mechanism by which CpG islands escape de novo methylation is sensitive to CpG density of adjacent sequences. These findings are of importance for the design of transgenes for controlled expression.