Inverse splicing of a discontinuous pre-mRNA intron generates a circular exon in a HeLa cell nuclear extract
Inverse splicing of a discontinuous pre-mRNA intron generates a circular exon in a HeLa cell nuclear extract
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DOI：
10.1093/nar/24.21.4152
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发表时间：
1996-11-01
影响因子：
14.9
通讯作者：
Wiebauer, K
中科院分区：
文献类型：
--
作者：
Braun, S;Domdey, H;Wiebauer, K
我们最近报道了第一个使用酿酒酵母全细胞提取物对真核细胞前mRNA内含子进行反向剪接的例子。在这种剪接反应过程中伴随的外显子环化产生了这样的假设，即最近在哺乳动物细胞中发现的环状RNA物种是通过反向剪接产生的。在这里，我们报道了在HeLa细胞核提取液中，通过人β-珠蛋白基因的第二内含子与前mRNA转录本的反向剪接反应形成环状外显子，其中两个内含子位于人工融合的单个外显子的两侧。我们的数据表明，哺乳动物的前mRNA剪接系统确实具有以相反的顺序对剪接位点进行比对的内在能力，并且这种比对之后可以进行完整的剪接反应，从而去除不连续的内含子序列。因此，我们提出了体内环状外显子的产生是5‘剪接位点与上游3’剪接位点配对后的反向剪接反应的结果，并且这一事件的发生频率受到其他竞争剪接位点的存在和强度的影响。
We have recently reported the first example of inverse splicing of a eukaryotic pre-mRNA intron using a whole cell extract from the yeast Saccharomyces cerevisiae. The concomitant circularization of the exon in the course of this splicing reaction gave rise to the hypothesis that the circular RNA species, which had been recently discovered in vivo in mammalian cells, were generated by inverse splicing, Here we report the formation of a circular exon in HeLa cell nuclear extracts by an inverse splicing reaction of the second intron of the human beta-globin gene from a pre-mRNA transcript in which the two intron halves flanked an artificially fused, single exon. Our data demonstrate that the mammalian pre-mRNA splicing system has indeed an intrinsic capability of aligning splice sites in reverse order and that this alignment can be followed by a complete splicing reaction, whereby the discontinuous intron sequences are removed. Thus we propose that circular exons in vivo arise as a result of an inverse splicing reaction following the pairing of a 5' splice site with an upstream 3' splice site and that the frequency of this event is influenced by the presence and strength of other, competing splice sites.