Seiwert S D, Heidmann S, Stuart K
Seattle Biomedical Research Institute, Washington, 98109, USA.
Cell. 1996 Mar 22;84(6):831-41. doi: 10.1016/s0092-8674(00)81062-4.
Deletion of uridylates from the 3'-most editing site of synthetic ATPase 6 pre-mRNA can be visualized directly by coincubation of a radiolabeled substrate RNA and a synthetic gRNA in 20S fractions of T.brucie mitochondrial lysates. Substrate RNA cleavage is gRNA directed and occurs 3' to the uridylates to be deleted. U residues appear to be sequentially removed from the 3' end of the 5' cleavage product prior to religation of the two pre-mRNA halves. gRNA/mRNA chimeric molecules are also produced. Time course experiments indicate that chimeras appear after cleavage intermediates and edited product. Furthermore, a mutant gRNA promotes formation of edited product but not detectable chimeras. Our results suggest a model for kinetoplastid RNA editing in which chimeric molecules are nonproductive end products of editing and not intermediates that serve as a repository for deleted U's.
通过将放射性标记的底物RNA与合成的引导RNA（gRNA）在布氏锥虫线粒体裂解物的20S组分中共同孵育，可以直接观察到从合成的ATP酶6前体mRNA的3'端最末端编辑位点删除尿苷酸的过程。底物RNA的切割是由gRNA引导的，并且发生在要删除的尿苷酸的3'端。在重新连接前体mRNA的两个半部分之前，U残基似乎是从5'切割产物的3'端依次去除的。还产生了gRNA/mRNA嵌合分子。时间进程实验表明，嵌合体在切割中间体和编辑产物之后出现。此外，一种突变的gRNA促进编辑产物的形成，但没有可检测到的嵌合体。我们的结果提出了一种动质体RNA编辑模型，其中嵌合分子是编辑的非生产性终产物，而不是作为删除的U的储存库的中间体。
