Spectroscopic observation of RNA chaperone activities of Hfq in post-transcriptional regulation by a small non-coding RNA.
Spectroscopic observation of RNA chaperone activities of Hfq in post-transcriptional regulation by a small non-coding RNA.
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DOI：
10.1093/nar/gkl1124
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发表时间：
2007
影响因子：
14.9
通讯作者：
Ha T
中科院分区：
文献类型：
--
作者：
Arluison V;Hohng S;Roy R;Pellegrini O;Régnier P;Ha T
Hfq蛋白对细菌中许多非编码小(S)RNA的功能是至关重要的，但Hfq促进sRNA功能的机制仍存在争议。我们建立了一种荧光共振能量转移分析方法来探索HfQ如何调节sRNA，DSRA，及其调控靶标mRNA，rpos之间的相互作用。相关的RNA片段被标记，从而可以实时监测分子内和分子间RNA结构的变化。我们的数据表明，HfQ促进了链交换反应，在该反应中，rpos的内部结构通过与DSRA配对而被取代，从而使mRNA的Shine-Dalgarno序列暴露。HFQ似乎通过诱导DSRA和预熔化的RPOS的快速结合以及帮助RPOS二级结构的缓慢破坏来进行链交换。出乎意料的是，HfQ还破坏了rpos和DSRA之间预先形成的复合体。虽然这在体内可能不是经常发生的事件，但这种融化活动可能与基于sRNA的调控的逆转有关。总体而言，我们的数据表明，Hfq不仅通过与DSRA和rpos快速结合来促进链交换，而且还具有RNA伴侣性质，促进动态RNA-RNA相互作用。
Hfq protein is vital for the function of many non-coding small (s)RNAs in bacteria but the mechanism by which Hfq facilitates the function of sRNA is still debated. We developed a fluorescence resonance energy transfer assay to probe how Hfq modulates the interaction between a sRNA, DsrA, and its regulatory target mRNA, rpoS. The relevant RNA fragments were labelled so that changes in intra- and intermolecular RNA structures can be monitored in real time. Our data show that Hfq promotes the strand exchange reaction in which the internal structure of rpoS is replaced by pairing with DsrA such that the Shine-Dalgarno sequence of the mRNA becomes exposed. Hfq appears to carry out strand exchange by inducing rapid association of DsrA and a premelted rpoS and by aiding in the slow disruption of the rpoS secondary structure. Unexpectedly, Hfq also disrupts a preformed complex between rpoS and DsrA. While it may not be a frequent event in vivo, this melting activity may have implications in the reversal of sRNA-based regulation. Overall, our data suggests that Hfq not only promotes strand exchange by binding rapidly to both DsrA and rpoS but also possesses RNA chaperoning properties that facilitates dynamic RNA–RNA interactions.