Chen S, Anderson K, Moore M J
Howard Hughes Medical Institute, Department of Biochemistry, MS 009, Brandeis University, 415 South Street, Waltham, MA 02454-9110, USA.
Proc Natl Acad Sci U S A. 2000 Jan 18;97(2):593-8. doi: 10.1073/pnas.97.2.593.
In most eukaryotic introns the 3' splice site is defined by a surprisingly short AG consensus found a variable distance downstream of the branch site. Exactly how the spliceosome determines which AG to use, however, is not well understood. Previously we showed that when the branch site and 3' splice site AG are supplied by separate RNA molecules, there is a strong preference for use of the 5'-most AG in the 3' splice site-containing RNA. Here we show that this apparent 5'-->3' directionality holds even when this RNA contains four tandem repeats of a 6-nt sequence containing AG. Exactly the same pattern of 3' splice site choice was observed when the same tandem repeats were incorporated into a full-length splicing substrate. When the 3' splice site AG is supplied by a separate RNA, that RNA must be linear with an unobstructed 5' end. Similarly, the branch-containing RNA must be truncated immediately 3' to the polypyrimidine tract. A model is presented that incorporates these observations and reconciles previously proposed mechanisms for 3' splice site selection.
在大多数真核生物内含子中，3'剪接位点由一个在分支位点下游可变距离处发现的惊人短的AG共有序列所界定。然而，剪接体究竟如何确定使用哪个AG，目前还不太清楚。此前我们表明，当分支位点和3'剪接位点AG由单独的RNA分子提供时，在含有3'剪接位点的RNA中强烈倾向于使用5'端最靠前的AG。在此我们表明，即使该RNA包含一个含有AG的6核苷酸序列的四个串联重复，这种明显的5'→3'方向性仍然成立。当相同的串联重复序列被整合到全长剪接底物中时，观察到完全相同的3'剪接位点选择模式。当3'剪接位点AG由单独的RNA提供时，该RNA必须是线性的且5'端无阻碍。同样，含分支的RNA必须在多嘧啶序列下游紧邻处被截断。本文提出了一个模型，该模型纳入了这些观察结果，并调和了先前提出的3'剪接位点选择机制。
