Mohr G, Lambowitz A M
Department of Molecular Genetics, Ohio State University, Columbus 43210.
Nature. 1991 Nov 14;354(6349):164-7. doi: 10.1038/354164a0.
Group I and II introns are mobile elements that propagate by insertion into different genes. Some introns of both types self-splice in vitro by transesterification reactions catalysed by the intron RNA. These transesterifications are reversible, and it has been suggested that reverse splicing followed by reverse transcription and recombination with genomic DNA may be a mechanism for intron transposition. In vivo the splicing of many, if not all, group I and II introns requires protein factors, which may facilitate correct folding of the intron RNAs. Here we show that the Neurospora mitochondrial large rRNA intron, a group I intron that is not self-splicing in vitro, undergoes reverse splicing in a reaction promoted by the CYT-18 protein, the Neurospora mitochondrial tyrosyl-tRNA synthetase, which is required for splicing the intron in vivo. In contrast to known RNA-catalysed reverse splicing reactions, this protein-assisted reverse splicing is sufficiently rapid to compete with forward splicing at low RNA concentrations under physiologically relevant conditions, including high GTP and low Mg2+ concentrations. Our results indicate that proteins that promote splicing could contribute to intron mobility by promoting reverse splicing in vivo.
I 类和 II 类内含子是通过插入不同基因进行传播的移动元件。这两类内含子中的一些在体外通过内含子 RNA 催化的酯交换反应进行自我剪接。这些酯交换反应是可逆的，有人提出，随后进行逆转录并与基因组 DNA 重组的反向剪接可能是内含子转座的一种机制。在体内，许多（如果不是全部的话）I 类和 II 类内含子的剪接需要蛋白质因子，这些蛋白质因子可能有助于内含子 RNA 的正确折叠。在这里，我们表明，粗糙脉孢菌线粒体大 rRNA 内含子，一种在体外不能自我剪接的 I 类内含子，在由 CYT-18 蛋白（粗糙脉孢菌线粒体酪氨酰-tRNA 合成酶，是该内含子在体内剪接所必需的）促进的反应中发生反向剪接。与已知的 RNA 催化的反向剪接反应不同，这种蛋白质辅助的反向剪接速度足够快，在生理相关条件下（包括高 GTP 和低 Mg2+浓度）的低 RNA 浓度下能够与正向剪接竞争。我们的结果表明，促进剪接的蛋白质可能通过在体内促进反向剪接来促进内含子的移动性。
