Department of Biochemistry, RNA Group, Université de Sherbrooke, Sherbrooke, Québec, Canada.
Nucleic Acids Res. 2018 Mar 16;46(5):2600-2612. doi: 10.1093/nar/gkx1287.
Small RNAs are key components of complex regulatory networks. These molecules can integrate multiple cellular signals to control specific target mRNAs. The recent development of high-throughput methods tremendously helped to characterize the full targetome of sRNAs. Using MS2-affinity purification coupled with RNA sequencing (MAPS) technology, we reveal the targetomes of two sRNAs, CyaR and RprA. Interestingly, both CyaR and RprA interact with the 5'-UTR of hdeD mRNA, which encodes an acid-resistance membrane protein. We demonstrate that CyaR classically binds to the RBS of hdeD, interfering with translational initiation. We identified an A/U-rich motif on hdeD, which is bound by the RNA chaperone Hfq. Our results indicate that binding of this motif by Hfq is required for CyaR-induced degradation of hdeD mRNA. Additional data suggest that two molecules of RprA must bind the 5'-UTR of hdeD to block translation initiation. Surprisingly, while both CyaR and RprA sRNAs bind to the same motif on hdeD mRNA, RprA solely acts at the translational level, leaving the target RNA intact. By interchanging the seed region of CyaR and RprA sRNAs, we also swap their regulatory behavior. These results suggest that slight changes in the seed region could modulate the regulation of target mRNAs.
小 RNA 是复杂调控网络的关键组成部分。这些分子可以整合多个细胞信号，以控制特定的靶 mRNA。高通量方法的最新发展极大地帮助了 sRNA 全靶标组的特征描述。我们使用 MS2 亲和纯化结合 RNA 测序（MAPS）技术，揭示了 CyaR 和 RprA 这两种 sRNA 的靶标组。有趣的是，CyaR 和 RprA 都与编码酸抗性膜蛋白的 hdeD mRNA 的 5'-UTR 相互作用。我们证明 CyaR 经典地结合 hdeD 的 RBS，干扰翻译起始。我们在 hdeD 上鉴定了一个富含 A/U 的基序，该基序被 RNA 伴侣 Hfq 结合。我们的结果表明，Hfq 结合该基序是 CyaR 诱导 hdeD mRNA 降解所必需的。额外的数据表明，两个 RprA 分子必须结合 hdeD 的 5'-UTR 以阻断翻译起始。令人惊讶的是，虽然 CyaR 和 RprA sRNA 都结合 hdeD mRNA 上的相同基序，但 RprA 仅在翻译水平起作用，使靶 RNA 保持完整。通过交换 CyaR 和 RprA sRNA 的种子区域，我们还交换了它们的调节行为。这些结果表明，种子区域的微小变化可以调节靶 mRNA 的调节。
