A highly active synthetic mammalian retrotransposon
A highly active synthetic mammalian retrotransposon
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DOI：
10.1038/nature02535
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发表时间：
2004-05-20
期刊：
影响因子：
64.8
通讯作者：
Boeke, JD
中科院分区：
文献类型：
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作者：
Han, JS;Boeke, JD
LINE-1 （L1）元件是逆转录转座子，包含哺乳动物基因组的大部分(1)。由于延伸缺陷，通过L1开放阅读框的转录效率低下(2)，抑制了L1 RNA和蛋白质、底物和反转录转位酶的强劲表达（3-5）。这种延伸缺陷可能控制着哺乳动物细胞L1转位的频率。在这里，我们报道通过合成寡核苷酸合成L1的开放阅读框来绕过这种转录缺陷，在不改变氨基酸序列的情况下改变24%的核酸序列。这种再合成导致稳态L1 RNA和蛋白质水平大大提高。值得注意的是，在已建立的反转录转位试验(4)中，当合成开放阅读框取代野生型开放阅读框时，转位水平增加了200多倍。这表明在L1编码区内可能不存在逆转录所需的大的、严格保守的顺式核酸序列。这些人工合成的反转录转座子也是迄今为止已知的最活跃的L1元件，具有作为操纵哺乳动物基因组的实用工具的潜力。
LINE-1 (L1) elements are retrotransposons that comprise large fractions of mammalian genomes(1). Transcription through L1 open reading frames is inefficient owing to an elongation defect(2), inhibiting the robust expression of L1 RNA and proteins, the substrate and enzyme(s) for retrotransposition(3-5). This elongation defect probably controls L1 transposition frequency in mammalian cells. Here we report bypassing this transcriptional defect by synthesizing the open reading frames of L1 from synthetic oligonucleotides, altering 24% of the nucleic acid sequence without changing the amino acid sequence. Such resynthesis led to greatly enhanced steady-state L1 RNA and protein levels. Remarkably, when the synthetic open reading frames were substituted for the wild-type open reading frames in an established retrotransposition assay(4), transposition levels increased more than 200-fold. This indicates that there are probably no large, rigidly conserved cis-acting nucleic acid sequences required for retrotransposition within L1 coding regions. These synthetic retrotransposons are also the most highly active L1 elements known so far and have potential as practical tools for manipulating mammalian genomes.