Liu Y, Huang T, MacMorris M, Blumenthal T
Department of Biochemistry and Molecular Genetics, University of Colorado Health Sciences Center, Denver 80262, USA.
RNA. 2001 Feb;7(2):176-81. doi: 10.1017/s1355838201002333.
About half of Caenorhabditis elegans genes have a 1-2 bp mismatch to the canonical AAUAAA hexamer that signals 3' end formation. One rare variant, AGUAAA, is found at the 3' end of the mai-1 gene, the first gene in an operon also containing gpd-2 and gpd-3. When we expressed this operon under heat shock control, 3' end formation dependent on the AGUAAA was very inefficient, but could be rescued by a single bp change to create a perfect AAUAAA. When AGUAAA was present, most 3' ends formed at a different site, 100 bp farther downstream, right at the gpd-2 trans-splice site. Surprisingly, 3' end formation at this site did not require any observable match to the AAUAAA consensus. It is possible that 3' end formation at this site occurs by a novel mechanism--trans-splicing-dependent cleavage--as deletion of the trans-splice site prevented 3' end formation here. Changing the AGUAAA to AAUAAA also influenced the trans-splicing process: with AGUAAA, most of the gpd-2 product was trans-spliced to SL1, rather than SL2, which is normally used at downstream operon trans-splice sites. However, with AAUAAA, SL2 trans-splicing of gpd-2 was increased. Our results imply that (1) the AAUAAA consensus controls 3' end formation frequency in C. elegans; (2) the AAUAAA is important in determining SL2 trans-splicing events more than 100 bp downstream; and (3) in some circumstances, 3' end formation may occur by a trans-splicing-dependent mechanism.
秀丽隐杆线虫约一半的基因与标志3'端形成的典型AAUAAA六聚体存在1-2个碱基对的错配。在mai-1基因的3'端发现了一种罕见变体AGUAAA，mai-1基因是一个操纵子中的第一个基因，该操纵子还包含gpd-2和gpd-3。当我们在热休克控制下表达这个操纵子时，依赖AGUAAA的3'端形成效率非常低，但通过单个碱基对的改变以产生完美的AAUAAA可以挽救这种情况。当存在AGUAAA时，大多数3'端在下游100 bp处的不同位点形成，正好在gpd-2反式剪接位点处。令人惊讶的是，该位点的3'端形成不需要与AAUAAA共有序列有任何可观察到的匹配。有可能该位点的3'端形成是通过一种新机制——反式剪接依赖性切割——发生的，因为反式剪接位点的缺失阻止了此处的3'端形成。将AGUAAA改变为AAUAAA也影响了反式剪接过程：对于AGUAAA，大多数gpd-2产物被反式剪接到SL1，而不是通常用于下游操纵子反式剪接位点的SL2。然而，对于AAUAAA，gpd-2的SL2反式剪接增加。我们的结果表明：（1）AAUAAA共有序列控制秀丽隐杆线虫中的3'端形成频率；（2）AAUAAA在确定下游100 bp以上的SL2反式剪接事件中比在确定下游100 bp以上的SL2反式剪接事件中更重要；（3）在某些情况下，3'端形成可能通过反式剪接依赖性机制发生。
