In vivo evidence for 5′→3′ exoribonuclease degradation of an unstable chloroplast mRNA
In vivo evidence for 5′→3′ exoribonuclease degradation of an unstable chloroplast mRNA
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DOI：
10.1046/j.1365-313x.1998.00016.x
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发表时间：
1998-01-01
期刊：
影响因子：
7.2
通讯作者：
Stern, DB
中科院分区：
文献类型：
--
作者：
Drager, RG;Girard-Bascou, J;Stern, DB
需要醋酸盐的莱茵衣藻核突变株F16含有突变mcd1-1，不能积累细胞色素b6/f复合体。Mcd1-1的主要缺陷是编码复合体IV亚单位的PETD mRNA不稳定。通过叶绿体转化引入的嵌合报告基因表明，在MCD(1-1)背景下PETD mRNA不稳定性的决定因素位于5‘非翻译区(UTR)。然而，当这个5‘UTR存在于双顺反子或嵌合转录本中其他序列的下游时，RNAs在mcd1-1背景中不再是不稳定的。综上所述，这些结果表明PETD 5‘UTR的5’端与MDC1产物相互作用。将多聚鸟苷序列插入到与报告基因融合的PETD 5‘UTR中，允许报告基因转录本在突变背景中积累。由于聚鸟苷形成了一种已知的阻碍核酸外切酶的结构，这些数据提供了体内证据，证明PETD mRNA可以被5‘-gt；3’外切核糖核酸酶活性降解。此外，这些数据支持一个模型，在该模型中，蛋白质与PETD 5‘UTR结合可以保护野生型细胞中的mRNA免受5’-gt；3‘降解。
The acetate-requiring Chlamydomonas reinhardtii nuclear mutant F16 harbors the mutation mcd1-1 and fails to accumulate the cytochrome b6/f complex. The primary defect of mcd1-1 was determined to be the instability of petD mRNA, which encodes subunit IV of the complex. Chimeric reporter genes introduced by chloroplast transformation demonstrated that the determinant of petD mRNA instability in the mcd(1-1) background is located in the 5' untranslated region (UTR). However, when this 5' UTR was present downstream of other sequences in dicistronic or chimeric transcripts, the RNAs were no longer destabilized in the mcd1-1 background. Together, these results suggest that the 5' end of the petD 5' UTR interacts with the MDC1 product. The insertion of a polyguanosine sequence into the petD 5' UTR fused to a reporter gene allowed accumulation of the reporter gene transcript in the mutant background. Since polyguanosine forms a structure that is known to impede exonucleases, these data provide in vivo evidence that petD mRNA can be degraded by 5' --> 3' exoribonuclease activity. Furthermore, the data support a model in which protein binding to the petD 5' UTR protects the mRNA from 5' --> 3' degradation in wild-type cells.