YEAST PRE-MESSENGER-RNA SPLICING EFFICIENCY DEPENDS ON CRITICAL SPACING REQUIREMENTS BETWEEN THE BRANCH POINT AND 3-' SPLICE SITE
YEAST PRE-MESSENGER-RNA SPLICING EFFICIENCY DEPENDS ON CRITICAL SPACING REQUIREMENTS BETWEEN THE BRANCH POINT AND 3-' SPLICE SITE
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DOI：
10.1002/j.1460-2075.1986.tb04317.x
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发表时间：
1986-05-01
期刊：
影响因子：
11.4
通讯作者：
ROSSI, JJ
中科院分区：
文献类型：
--
作者：
CELLINI, A;FELDER, E;ROSSI, JJ
在酵母中，5”和3”剪接连接和内部分支受体位点（TACTAAC盒）是高度保守的内含子元件。对突变体的分析已经证明了这些元素在剪接过程中的重要性。在本报告中，我们通过三种不同的分析方法(剪接依赖性。β。-半乳糖苷酶表达，体外剪接分析和体内RNA分析)，至少两个元件（TACTAAC和3”剪接信号）也必须满足一定的间距要求，以允许有效的剪接发生。特别是，3‘’剪接位点与2''-5‘’分支位点的间距是决定剪接、内含子释放和外显子-外显子连接最终反应完成效率的关键因素。虽然该区域内的插入对剪接的第一反应（5‘’切割和2''-5‘’分支的形成）几乎没有影响，但它们极大地影响了最终反应的效率。相比之下，这两个位点之间的15个碱基缺失对剪接效率没有可检测到的影响。我们还发现，在分枝位点下游的所有酵母菌序列都被大肠杆菌序列所取代的转录本中，尽管效率不高，但5‘’切割和分支形成可以发生。我们从这些研究中得出结论，在酵母中，5‘’和3‘’的剪接位点是彼此独立识别的，但总是与TACTAAC信号结合在一起。
In the yeast Saccharomyces cerevisiae the 5'' and 3'' splice junctions and the internal branch acceptor site (TACTAAC box) are highly conserved intron elements. Analyses of mutants have demonstrated the importance of each of these elements in the splicing process. In the present report we show by three different analytical approaches (splicing-dependent .beta.-galactosidase expression, in vitro splicing assays and in vivo RNA analyses) that at least two of these elements (the TACTAAC and 3'' splice signals) also have to fulfill certain spacing requirements to allow efficient splicing to occur. In particular, the spacing of the 3'' splice site from the 2''-5'' branch site is a critical factor in determining the efficiency for completion of the final reactions of splicing, intron release and exon-exon joining. Whereas insertions within this region have little or no effect on the first reactions in splicing (the 5'' cleavage and 2''-5'' branch formation), they dramatically affect the efficiency of the final reactions. In contrast, a 15-base deletion between these two sites has no detectable effect on splicing efficiency. We also show that the 5'' cleavage and branch formation can take place, albeit inefficiently, in transcripts in which all of the yeast sequences downstream of the branch site have been replaced by Escherichia coli sequences. We conclude from these studies that, in yeast, the 5'' and 3'' splice sites are recognized independently from one another, but always in conjunction with the TACTAAC signal.