Center for Infectious Diseases and Vaccinology, Biodesign Institute, and Arizona State University, Tempe, Arizona 85287-4501; School of Life Sciences, Arizona State University, Tempe, Arizona 85287-4501.
Center for Infectious Diseases and Vaccinology, Biodesign Institute, and Arizona State University, Tempe, Arizona 85287-4501.
J Biol Chem. 2014 Apr 18;289(16):11353-11366. doi: 10.1074/jbc.M113.517516. Epub 2014 Mar 4.
Riboswitches are a class of cis-acting regulatory RNAs normally characterized from the 5'-UTR of bacterial transcripts that bind a specific ligand to regulate expression of associated genes by forming alternative conformations. Here, we present a riboswitch that contributes to transcriptional regulation through sensing Mn(2+) in Salmonella typhimurium. We characterized a 5'-UTR (UTR1) from the mntH locus encoding a Mn(2+) transporter, which forms a Rho-independent terminator to implement transcription termination with a high Mn(2+) selectivity both in vivo and in vitro. Nucleotide substitutions that cause disruption of the terminator interfere with the regulatory function of UTR1. RNA probing analyses outlined a specific UTR1 conformation that favors the terminator structure in Mn(2+)-replete condition. Switch sequence GCUAUG can alternatively base pair duplicated hexanucleotide CAUAGC to form either a pseudoknot or terminator stem. Mn(2+), but not Mg(2+), and Ca(2+), can enhance cleavage at specific nucleotides in UTR1. We conclude that UTR1 is a riboswitch that senses cytoplasmic Mn(2+) and therefore participates in Mn(2+)-responsive mntH regulation in Salmonella. This riboswitch domain is also conserved in several Gram-negative enteric bacteria, indicating that this Mn(2+)-responsive mechanism could have broader implications in bacterial gene expression. Additionally, a high level of cytoplasmic Mn(2+) can down-regulate transcription of the Salmonella Mg(2+) transporter mgtA locus in a Mg(2+) riboswitch-dependent manner. On the other hand, these two types of cation riboswitches do not share similarity at the primary or secondary structural levels. Taken together, characterization of Mn(2+)-responsive riboswitches should expand the scope of RNA regulatory elements in response to inorganic ions.
Riboswitches 是一类顺式作用调节 RNA，通常从细菌转录本的 5'-UTR 中鉴定出来，通过形成不同的构象结合特定的配体来调节相关基因的表达。在这里，我们介绍了一种在鼠伤寒沙门氏菌中通过感应 Mn(2+)参与转录调控的 riboswitch。我们对编码 Mn(2+)转运蛋白的 mntH 基因座的 5'-UTR（UTR1）进行了特征描述，该 UTR1 形成 Rho 独立终止子，在体内和体外都具有高 Mn(2+)选择性实现转录终止。导致终止子破坏的核苷酸取代会干扰 UTR1 的调节功能。RNA 探测分析概述了一种特定的 UTR1 构象，该构象在 Mn(2+)充足的条件下有利于终止子结构。开关序列 GCUAUG 可以替代碱基配对重复的六核苷酸 CAUAGC，形成假结或终止子茎。只有 Mn(2+)，而不是 Mg(2+)和 Ca(2+)，可以增强 UTR1 中特定核苷酸的切割。我们得出结论，UTR1 是一种感应细胞质 Mn(2+)的 riboswitch，因此参与了鼠伤寒沙门氏菌中 Mn(2+)响应的 mntH 调节。这个 riboswitch 结构域在几种革兰氏阴性肠杆菌中也保守，这表明这种 Mn(2+)响应机制在细菌基因表达中可能具有更广泛的意义。此外，细胞质中高水平的 Mn(2+)可以以 Mg(2+) riboswitch 依赖的方式下调沙门氏菌 Mg(2+)转运蛋白 mgtA 基因座的转录。另一方面，这两种类型的阳离子 riboswitches 在一级或二级结构水平上没有相似性。综上所述，Mn(2+)响应 riboswitch 的特征描述应该扩大对无机离子反应的 RNA 调节元件的范围。
