The rpoS mRNA leader recruits Hfq to facilitate annealing with DsrA sRNA
The rpoS mRNA leader recruits Hfq to facilitate annealing with DsrA sRNA
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DOI：
10.1261/rna.1110608
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发表时间：
2008-09-01
期刊：
影响因子：
4.5
通讯作者：
Woodson, Sarah A.
中科院分区：
文献类型：
--
作者：
Soper, Toby J.;Woodson, Sarah A.
小非编码 RNA (sRNA) 与 Sm 样 RNA 结合蛋白 Hfq 一起调节细菌对环境应激的反应。 DsrA sRNA 通过与 rpoS mRNA 的 5' 前导序列碱基配对并打开抑制翻译起始的茎环，刺激大肠杆菌中 RpoS 应激反应因子的翻译。我们报告说，该茎环上游的 rpoS 前导序列大大增加了 rpoS mRNA 对 Hfq 和 DsrA 的敏感性。天然凝胶迁移率变化测定表明，Hfq 使 DsrA 与完整 576 nt rpoS 先导序列的结合率提高了 50 倍。相比之下，与仅包含阻遏物茎环的 138 nt RNA 的碱基配对仅加速两倍。缺失和诱变实验表明，对 Hfq 的敏感性需要上游 AAYAA 序列。足够长以包含该序列的前导序列紧密结合 Hfq 并与 Hfq 和 DsrA 形成稳定的三元复合物。提出了一种模型，其中 Hfq 通过结合两个 RNA 将 DsrA 募集到 rpoS mRNA，释放 mRNA 中的自我抑制结构。一旦 DsrA 和 rpoS mRNA 之间的碱基配对建立，Hfq 和 mRNA 之间的相互作用可以通过从 sRNA 中去除 Hfq 来稳定 RNA 复合物。
Small noncoding RNAs (sRNAs) regulate the response of bacteria to environmental stress in conjunction with the Sm-like RNA binding protein Hfq. DsrA sRNA stimulates translation of the RpoS stress response factor in Escherichia coli by base-pairing with the 5' leader of the rpoS mRNA and opening a stem-loop that represses translation initiation. We report that rpoS leader sequences upstream of this stem-loop greatly increase the sensitivity of rpoS mRNA to Hfq and DsrA. Native gel mobility shift assays show that Hfq increases the rate of DsrA binding to the full 576 nt rpoS leader as much as 50-fold. By contrast, base-pairing with a 138-nt RNA containing just the repressor stem-loop is accelerated only twofold. Deletion and mutagenesis experiments showed that sensitivity to Hfq requires an upstream AAYAA sequence. Leaders long enough to contain this sequence bind Hfq tightly and form stable ternary complexes with Hfq and DsrA. A model is proposed in which Hfq recruits DsrA to the rpoS mRNA by binding both RNAs, releasing the self-repressing structure in the mRNA. Once base-pairing between DsrA and rpoS mRNA is established, interactions between Hfq and the mRNA may stabilize the RNA complex by removing Hfq from the sRNA.