Upstream Flanking Sequence Assists Folding of an RNA Thermometer.
Upstream Flanking Sequence Assists Folding of an RNA Thermometer.
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DOI：
10.1016/j.jmb.2022.167786
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发表时间：
2022-09-30
影响因子：
5.6
通讯作者：
Bevilacqua, Philip C.
中科院分区：
文献类型：
--
作者：
Jolley, Elizabeth A.;Bormes, Kathryn M.;Bevilacqua, Philip C.
细菌中的许多热休克基因是通过一类称为 RNA 温度计 (RNAT) 的温度敏感茎环 (SL) RNA 来调节的。研究最广泛的 RNAT 之一是与热休克蛋白表达相关的热休克表达抑制 (ROSE) 元件。 RNAT 位于 5'UTR，其上游包含一到三个辅助发夹。在此，我们探讨了这些上游 SL 在 RNAT 折叠和功能中的作用。日本慢生根瘤菌 (Bradyrhizobium japonicum) 是一种固氮细菌，可在土壤中经历较宽的温度范围，并含有 ROSE 元素，每个元素都有多个上游 SL。日本芽孢杆菌热休克蛋白 A (hspA) 信使 (mRNA) 的 5’UTR 具有复杂的二级结构，包含 RNAT SL 上游的三个 SL。虽然 hspA RNAT 本身的结构功能研究已有报道，但尚不清楚这些辅助 SL 是否有助于 ROSE 元件的温度传感功能。在此，我们证明全长（FL）序列具有多个熔解转变，表明ROSE元件以非二态方式展开。上游 SL 比 RNAT 本身更稳定，紧邻 RNAT 上游的 SL 中碱基配对中断的变体对 RNAT 的解链几乎没有影响。基于这些结果和 ROSE 元件共转录折叠的建模，我们提出上游 SL 的功能是稳定转录物并帮助 RNAT 的正确折叠和动力学。
Many heat shock genes in bacteria are regulated through a class of temperature-sensitive stem-loop (SL) RNAs called RNA thermometers (RNATs). One of the most widely studied RNATs is the Repression Of heat Shock Expression (ROSE) element associated with expression of heat shock proteins. Located in the 5’UTR, the RNAT contains one to three auxiliary hairpins upstream of it. Herein, we address roles of these upstream SLs in the folding and function of an RNAT. Bradyrhizobium japonicum is a nitrogen-fixing bacterium that experiences a wide range of temperatures in the soil and contains ROSE elements, each having multiple upstream SLs. The 5’UTR of the messenger (mRNA) for heat shock protein A (hspA) in B. japonicum has an intricate secondary structure containing three SLs upstream of the RNAT SL. While structure-function studies of the hspA RNAT itself have been reported, it has been unclear if these auxiliary SLs contribute to the temperature-sensing function of the ROSE elements. Herein, we show that the full length (FL) sequence has several melting transitions indicating that the ROSE element unfolds in a non-two-state manner. The upstream SLs are more stable than the RNAT itself, and a variant with disrupted base pairing in the SL immediately upstream of the RNAT has little influence on the melting of the RNAT. On the basis of these results and modeling of the co-transcriptional folding of the ROSE element, we propose that the upstream SLs function to stabilize the transcript and aid proper folding and dynamics of the RNAT.
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