Decker C J, Parker R
Department of Molecular and Cellular Biology, University of Arizona, Tucson 85721.
Genes Dev. 1993 Aug;7(8):1632-43. doi: 10.1101/gad.7.8.1632.
To determine pathways of mRNA turnover in yeast, we have followed the poly(A) tail removal and degradation of a pulse of newly synthesized transcripts from four different genes. Before decay of both stable and unstable mRNAs initiated, there was a temporal lag during which the poly(A) tail was deadenylated to an oligo(A) length. Altering the deadenylation rate of an mRNA led to a corresponding change in the length of this lag. The rate of deadenylation and the stability of the oligo(A) species varied between mRNAs, explaining the differences in mRNA half-lives. To examine how the transcript body was degraded following deadenylation, we used the strategy of inserting strong RNA secondary structures, which can slow exonucleolytic digestion and thereby trap decay intermediates, into the 3' UTR of mRNAs. Fragments lacking the 5' portion of two different mRNAs accumulated after deadenylation as full-length mRNA levels decreased. Therefore, these results define an mRNA decay pathway in which deadenylation leads to either internal cleavage or decapping followed by 5'-->3' exonucleolytic degradation of the mRNA.
为了确定酵母中mRNA周转的途径，我们追踪了来自四个不同基因的新合成转录本脉冲的多聚腺苷酸（poly(A)）尾去除和降解过程。在稳定和不稳定mRNA开始降解之前，存在一个时间延迟，在此期间poly(A)尾被去腺苷酸化至寡聚腺苷酸长度。改变mRNA的去腺苷酸化速率会导致该延迟长度相应改变。不同mRNA之间，去腺苷酸化速率和寡聚腺苷酸种类的稳定性有所不同，这解释了mRNA半衰期的差异。为了研究去腺苷酸化后转录本主体如何降解，我们采用了将能够减缓核酸外切消化从而捕获降解中间体的强RNA二级结构插入mRNA的3'非翻译区（UTR）的策略。随着全长mRNA水平下降，缺乏两种不同mRNA 5'部分的片段在去腺苷酸化后积累。因此，这些结果定义了一种mRNA降解途径，其中去腺苷酸化导致mRNA内部切割或脱帽，随后进行5'→3'核酸外切降解。
