Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA.
J Mol Biol. 2011 Jan 7;405(1):3-17. doi: 10.1016/j.jmb.2010.09.064. Epub 2010 Oct 8.
To produce mature ribosomal RNAs (rRNAs), polycistronic rRNA transcripts are cleaved in an ordered series of events. We have uncovered the molecular basis for the ordering of two essential cleavage steps at the 3'-end of 18S rRNA. Using in vitro and in vivo structure probing, RNA binding and cleavage experiments, and yeast genetics, we demonstrate that a conserved RNA sequence in the spacer region between the 18S and 5.8S rRNAs base-pairs with the decoding site of 18S rRNA in early assembly intermediates. Nucleolar cleavage at site A(2) excises this sequence element, leading to a conformational switch in pre-18S rRNA, by which the ribosomal decoding site is formed. This conformational switch positions the nuclease Nob1 for cytoplasmic cleavage at the 3'-end of 18S rRNA and is required for the final maturation step of 18S rRNA in vivo and in vitro. More generally, our data show that the intrinsic ability of RNA to form stable structural switches is exploited to order and regulate RNA-dependent biological processes.
为了产生成熟的核糖体 RNA（rRNA），多顺反子 rRNA 转录物会按照一系列有序事件进行切割。我们已经揭示了 18S rRNA 3'端两个关键切割步骤的排序的分子基础。通过体外和体内结构探测、RNA 结合和切割实验以及酵母遗传学，我们证明了在 18S 和 5.8S rRNA 之间的间隔区的保守 RNA 序列与 18S rRNA 的解码位点在早期组装中间体中进行碱基配对。核仁切割位点 A(2) 切除这个序列元件，导致 pre-18S rRNA 发生构象转换，从而形成核糖体的解码位点。这种构象转换使核酶 Nob1 能够在细胞质中对 18S rRNA 的 3'端进行切割，这是 18S rRNA 在体内和体外进行最终成熟步骤所必需的。更普遍地说，我们的数据表明，RNA 形成稳定结构开关的内在能力被利用来对 RNA 依赖的生物过程进行排序和调节。
