Factors influencing alternative splice site utilization in vivo.
Factors influencing alternative splice site utilization in vivo.
复制标题
影响体内选择性剪接位点利用的因素。
DOI：
10.1128/mcb.7.2.738-748.1987
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发表时间：
1987
影响因子：
5.3
通讯作者：
Manley,JL
中科院分区：
文献类型：
--
作者：
Fu,XY;Manley,JL
为了研究影响备选前mrna剪接途径选择的因素，我们将表达野生型或突变的猿猴病毒40 （SV40）早期区域的质粒引入组织培养细胞，然后测量产生的小t和大t rna的数量。控制剪接位点选择的一个重要因素被发现是在产生small-t mRNA时去除的内含子的大小；该内含子的扩增（从66个核苷酸扩增到77个或更多核苷酸）导致相对于大t mRNA产生的小t mRNA数量大幅增加。这表明，在SV40早期pre-mRNA加工的正常过程中，由于后者内含子的尺寸较小，大t剪接相对于小t剪接具有竞争优势。通过分析包含剪接位点重复的突变的影响，还确定了前体mrna的几个可以影响剪接位点选择的附加特征。这包括竞争性的5 '剪接位点的优势和剪接位点在mrna前体中的相对位置。最后，我们发现人293细胞中小t mRNA与大t mRNA的比例是HeLa细胞或其他哺乳动物细胞类型的10至15倍。这些结果表明细胞特异性反式作用因子的存在可以显著改变pre-mRNA的剪接位点选择模式。
To study factors that influence the choice of alternative pre-mRNA splicing pathways, we introduced plasmids expressing either wild-type or mutated simian virus 40 (SV40) early regions into tissue culture cells and then measured the quantities of small-t and large-T RNAs produced. One important element controlling splice site selection was found to be the size of the intron removed in the production of small-t mRNA; expansion of this intron (from 66 to 77 or more nucleotides) resulted in a substantial increase in the amount of small-t mRNA produced relative to large-T mRNA. This suggests that in the normal course of SV40 early pre-mRNA processing, large-T splicing is at a competitive advantage relative to small-t splicing because of the small size of the latter intron. Several additional features of the pre-mRNA that can influence splice site selection were also identified by analyzing the effects of mutations containing splice site duplications. These include the strengths of competing 5′ splice sites and the relative positions of splice sites in the pre-mRNA. Finally, we showed that the ratio of small-t to large-T mRNA was 10 to 15-fold greater in human 293 cells than in HeLa cells or other mammalian cell types. These results suggest the existence of cell-specifictrans-acting factors that can dramatically alter the pattern of splice site selection in a pre-mRNA.