Niwa M, Rose S D, Berget S M
Marrs McClean Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030.
Genes Dev. 1990 Sep;4(9):1552-9. doi: 10.1101/gad.4.9.1552.
The majority of vertebrate pre-mRNAs are both spliced and polyadenylated. To investigate the mechanism whereby processing factors recognize last exons containing both splicing and polyadenylation consensus elements, chimeric precursor RNAs containing a single intron and a poly(A) site were constructed and assayed for in vitro splicing and polyadenylation. Chimeric RNAs underwent splicing and polyadenylation. Both reactions occurred in a single RNA. The presence of an intron enhanced the rate of polyadenylation at a downstream poly(A) site. The extent of stimulation varied from two- to fivefold, depending on the magnesium concentration. Maximal stimulation of polyadenylation by an upstream intron required a 3' splice site but not a 5' splice site, suggesting that the structure of the terminal exon was more important than the presence of a complete upstream intron. We suggest that splicing and polyadenylation factors interact to recognize terminal, poly(A) site-containing exons. Such interaction may explain why all known intron-containing eukaryotic pre-mRNAs generate their 3' ends by polyadenylation.
大多数脊椎动物的前体mRNA既会发生剪接又会进行多聚腺苷酸化。为了研究加工因子识别同时包含剪接和多聚腺苷酸化共有元件的最后一个外显子的机制，构建了含有单个内含子和一个多聚腺苷酸化位点的嵌合前体RNA，并对其进行体外剪接和多聚腺苷酸化检测。嵌合RNA发生了剪接和多聚腺苷酸化。这两个反应都发生在单个RNA上。内含子的存在提高了下游多聚腺苷酸化位点的多聚腺苷酸化速率。刺激程度在两倍到五倍之间变化，这取决于镁离子浓度。上游内含子对多聚腺苷酸化的最大刺激需要一个3'剪接位点，但不需要5'剪接位点，这表明末端外显子的结构比完整上游内含子的存在更重要。我们认为剪接因子和多聚腺苷酸化因子相互作用以识别含多聚腺苷酸化位点的末端外显子。这种相互作用可能解释了为什么所有已知的含内含子的真核前体mRNA都通过多聚腺苷酸化产生其3'末端。
