Ribonucleoprotein particle formation is necessary but not sufficient for LINE-1 retrotransposition
Ribonucleoprotein particle formation is necessary but not sufficient for LINE-1 retrotransposition
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DOI：
10.1093/hmg/ddi354
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发表时间：
2005-11-01
影响因子：
3.5
通讯作者：
Moran, JV
中科院分区：
文献类型：
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作者：
Kulpa, DA;Moran, JV
长散布元件（LINE-1或L1）是丰富的非LTR逆转录转座子，通过靶位点引发的逆转录通过RNA中间体进行动员。L1编码的蛋白质（ORF 1 p和ORF 2 p）优先与其编码的转录物结合形成核糖核蛋白颗粒（RNP），其是提议的反转录转座中间体。在这里，我们已经使用表位标记来区分由工程L1编码的蛋白质与由内源性表达的L1编码的蛋白质。我们证明，在ORF 1 p的羧基末端含有表位标签的L1仍然具有反转录转座能力，并且标记的ORF 1 p及其编码RNA定位于细胞质RNP。我们还鉴定了两类ORF 1 p突变体，一类严重减少RNP形成并阻断反转录转座，另一类允许RNP形成但使反转录转座减少100倍。因此，这些数据表明，RNP的形成是重要的，但不足以为L1反转录转座，并建议ORF 1 p也可能在下游步骤中的L1反转录转座途径的功能。
Long interspersed elements (LINE-1s or L1s) are abundant non-LTR retrotransposons that mobilize through an RNA intermediate by target site primed reverse transcription. The L1-encoded proteins (ORF1p and ORF2p) preferentially associate with their encoding transcript to form a ribonucleoprotein particle (RNP), which is a proposed retrotransposition intermediate. Here, we have used epitope tagging to discriminate the proteins encoded by engineered L1s from those encoded by endogenously expressed L1s. We demonstrate that an L1 containing an epitope tag at the carboxyl terminus of ORF1p remains retrotransposition-competent and that tagged ORF1p and its encoding RNA localize to cytoplasmic RNPs. We also identified two classes of ORF1p mutants, one that severely decreased RNP formation and blocked retrotransposition, and another that allows RNP formation but reduces retrotransposition by 100-fold. Thus, these data indicate that RNP formation is important but not sufficient for L1 retrotransposition and suggest that ORF1p also may function at downstream steps in the L1 retrotransposition pathway.