Essential motifs in the 3′ untranslated region required for retrotransposition and the precise start of reverse transcription in non-long-terminal- repeat retrotransposon SART1
Essential motifs in the 3′ untranslated region required for retrotransposition and the precise start of reverse transcription in non-long-terminal- repeat retrotransposon SART1
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DOI：
10.1128/mcb.24.18.7902-7913.2004
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发表时间：
2004-09-01
影响因子：
5.3
通讯作者：
Fujiwara, H
中科院分区：
文献类型：
--
作者：
Osanai, M;Takahashi, H;Fujiwara, H
非长端重复（Non-long-terminal-repeat, non-LTR）逆转录转座子通过从3'端逆转录mRNA来扩增其拷贝，但逆转录的初始过程尚不清楚。我们已经证明，蚕的端粒特异性非ltr反转录转座子SART1需要3‘非翻译区（3’ UTR）进行反转录转座子。通过体内逆转录试验，我们在3' UTR中发现了几个新的基序，这些基序参与了精确和有效的逆转录。在3' UTR的461个核苷酸（nt）中，从nt 163到nt 295的中心区域是SART1逆转录所必需的。在RNA中为SART1 3' UTR形成的五个假定的茎环中，第二个茎环（nt 159 - 221）包含在该区域。3‘ UTR和poly(A)链中3’区（nt 296 - 461）的缺失导致逆转录减少和不准确，这主要是从第二个茎环下游的几个端粒重复样GGUU序列开始的。这些结果表明，3' UTR中的短端粒重复序列类似于（TTAGG）(n)重复序列的底部链。我们还证明，当绿色荧光蛋白（GFP）编码区与SART1 3' UTR融合时，绿色荧光蛋白（GFP）的mRNA可以反转录成端粒重复序列，而SART1开放阅读框蛋白则以反式方式提供。
Non-long-terminal-repeat (non-LTR) retrotransposons amplify their copies by reverse transcribing mRNA from the 3' end, but the initial processes of reverse transcription are still unclear. We have shown that a telomere-specific non-LTR retrotransposon of the silkworm, SART1, requires the 3' untranslated region (3' UTR) for retrotransposition. With an in vivo retrotransposition assay, we identified several novel motifs within the 3' UTR involved in precise and efficient reverse transcription. Of 461 nucleotides (nt) of the 3' UTR, the central region, from nt 163 to nt 295, was essential for SART1 retrotransposition. Of five putative stem-loops formed in RNA for the SART1 3' UTR, the second stem-loop (nt 159 to 221) is included in this region. Loss of the 3' region (nt 296 to 461) in the 3' UTR and the poly(A) tract resulted in decreased and inaccurate reverse transcription, which starts mostly from several telomeric repeat-like GGUU sequences just downstream of the second stem-loop. These results suggest that short telomeric repeat-like sequences in the 3' UTR anneal to the bottom strand of (TTAGG)(n) repeats. We also demonstrated that the mRNA for green fluorescent protein (GFP) could be retrotransposed into telomeric repeats when the GFP coding region is fused with the SART1 3' UTR and SART1 open reading frame proteins are supplied in trans.