Walsh A P, Tock M R, Mallen M H, Kaberdin V R, von Gabain A, McDowall K J
Astbury Centre for Structural Molecular Biology, Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
Nucleic Acids Res. 2001 May 1;29(9):1864-71. doi: 10.1093/nar/29.9.1864.
RNase E initiates the decay of Escherichia coli RNAs by cutting them internally near their 5'-end and is a component of the RNA degradosome complex, which also contains the 3'-exonuclease PNPASE: Recently, RNase E has been shown to be able to remove poly(A) tails by what has been described as an exonucleolytic process that can be blocked by the presence of a phosphate group on the 3'-end of the RNA. We show here, however, that poly(A) tail removal by RNase E is in fact an endonucleolytic process that is regulated by the phosphorylation status at the 5'- but not the 3'-end of RNA. The rate of poly(A) tail removal by RNase E was found to be 30-fold greater when the 5'-terminus of RNA substrates was converted from a triphosphate to monophosphate group. This finding prompted us to re-analyse the contributions of the ribonucleolytic activities within the degradosome to 3' attack since previous studies had only used substrates that had a triphosphate group on their 5'-end. Our results indicate that RNase E associated with the degradosome may contribute to the removal of poly(A) tails from 5'-monophosphorylated RNAs, but this is only likely to be significant should their attack by PNPase be blocked.
核糖核酸酶E通过在大肠杆菌RNA的5'端附近进行内部切割来启动其降解，并且是RNA降解体复合物的一个组成部分，该复合物还包含3'核酸外切酶PNPASE。最近，已证明核糖核酸酶E能够通过一种被描述为核酸外切过程的方式去除聚腺苷酸尾巴，该过程可被RNA 3'端存在的磷酸基团所阻断。然而，我们在此表明，核糖核酸酶E去除聚腺苷酸尾巴实际上是一个核酸内切过程，它受RNA 5'端而非3'端的磷酸化状态调控。当RNA底物的5'末端从三磷酸基团转变为单磷酸基团时，发现核糖核酸酶E去除聚腺苷酸尾巴的速率提高了30倍。这一发现促使我们重新分析降解体中核糖核酸酶活性对3'端攻击的贡献，因为之前的研究仅使用了在其5'端具有三磷酸基团的底物。我们的结果表明，与降解体相关的核糖核酸酶E可能有助于从5'单磷酸化RNA上去除聚腺苷酸尾巴，但只有在PNPASE对其的攻击被阻断时，这才可能具有显著意义。
