Makino Shin-ichi, Sawasaki Tatsuya, Tozawa Yuzuru, Endo Yaeta, Takai Kazuyuki
Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan.
Biochem Biophys Res Commun. 2006 Sep 8;347(4):1080-7. doi: 10.1016/j.bbrc.2006.07.011. Epub 2006 Jul 14.
Cell extracts from wheat embryos have been widely used for mRNA-directed protein production. Here, we found that a significant fraction of exogenous linear RNAs are circularized in a wheat embryo extract. The circularization was seen only in uncapped RNAs. The amount of the circular species reached around 1% of the initial RNA and increased along with an increase in the initial concentration more than proportionally. The circular RNAs were stable but unable to be translated in the extract. The circularization was competitively inhibited in the presence of a known substrate of a wheat embryo RNA ligase. Thus, we cloned the RNA ligase cDNAs. Three isoform sequences were homologous to the other plant RNA ligases. An addition of a cell-free synthesized wheat RNA ligase abolished the inhibition, which indicates a participation of its activity in the circularization. A possible role in RNA metabolism, RNA silencing in particular, is discussed.
小麦胚细胞提取物已被广泛用于mRNA指导的蛋白质生产。在此，我们发现相当一部分外源线性RNA在小麦胚提取物中发生了环化。这种环化仅在无帽RNA中可见。环状RNA的量达到初始RNA的约1%，并随着初始浓度的增加而超比例增加。环状RNA是稳定的，但在提取物中无法翻译。在存在小麦胚RNA连接酶已知底物的情况下，环化受到竞争性抑制。因此，我们克隆了RNA连接酶的cDNA。三个同工型序列与其他植物RNA连接酶同源。添加无细胞合成的小麦RNA连接酶消除了抑制作用，这表明其活性参与了环化过程。文中还讨论了其在RNA代谢，特别是RNA沉默中的可能作用。
