A Tale of Two Transitions: The Unfolding Mechanism of the prfA RNA Thermosensor
A Tale of Two Transitions: The Unfolding Mechanism of the prfA RNA Thermosensor
复制标题
两次转变的故事：prfA RNA 热传感器的展开机制
DOI：
10.1021/acs.biochem.0c00588
复制
发表时间：
2020
期刊：
影响因子：
2.9
通讯作者：
Keane, Sarah C.
中科院分区：
文献类型：
--
作者：
Zhang, Huaqun;Hall, Ian;Nissley, Amos J.;Abdallah, Kyrillos;Keane, Sarah C.
RNA 热传感器 (RNAT) 存在于一些细菌信使 RNA (mRNA) 的 5' 非翻译区 (UTR)，以温度依赖性方式控制下游基因的翻译。在单核细胞增生李斯特氏菌中，关键转录因子 PrfA 的表达是由其 5' UTR 中的 RNAT 介导的。 PrfA 充当 L 毒力的主要调节因子。单核细胞增多症，控制许多毒力因子的表达。 PrfA 通过其 RNAT 元件进行温度调节表达，是细菌成功入侵宿主的信号。在结构上，prfA RNAT 与已知的 RNAT 家族几乎没有相似之处，之前的研究表明 prfA RNAT 在较窄的温度范围内具有高度响应性。在这里，我们进行了全面的突变和热力学分析，以确定温度敏感性的分子决定因素。我们提供的证据支持这样的观点，即 prfA RNAT 的展开与 cssA（一种充分表征的 RNAT）的展开不同，表明这些 RNAT 通过不同的机制发挥作用。我们的数据表明，prfA RNAT 的解折叠发生在两个不同的事件中，并且内部环在介导 RNAT 解折叠的协同性中发挥着重要作用。我们进一步证明，核糖体结合位点 (RBS) 远端的区域不仅有助于 RNAT 结构稳定性，而且还影响下游信息的翻译。我们的集体结果提供了将 prfA RNAT 结构的热稳定性、展开能量学和翻译控制联系起来的见解。
RNA thermosensors (RNATs), found in the 5′ untranslated region (UTR) of some bacterial messenger RNAs (mRNAs), control the translation of the downstream gene in a temperature-dependent manner. InListeria monocytogenes, the expression of a key transcription factor, PrfA, is mediated by an RNAT in its 5′ UTR. PrfA functions as a master regulator of virulence inL. monocytogenes, controlling the expression of many virulence factors. The temperature-regulated expression of PrfA by its RNAT element serves as a signal of successful host invasion for the bacteria. Structurally, the prfA RNAT bears little resemblance to known families of RNATs, and prior studies demonstrated that the prfA RNAT is highly responsive over a narrow temperature range. Herein, we have undertaken a comprehensive mutational and thermodynamic analysis to ascertain the molecular determinants of temperature sensitivity. We provide evidence to support the idea that the prfA RNAT unfolding is different from that of cssA, a well-characterized RNAT, suggesting that these RNATs function via distinct mechanisms. Our data show that the unfolding of the prfA RNAT occurs in two distinct events and that the internal loops play an important role in mediating the cooperativity of RNAT unfolding. We further demonstrated that regions distal to the ribosome binding site (RBS) not only contribute to RNAT structural stability but also impact translation of the downstream message. Our collective results provide insight connecting the thermal stability of the prfA RNAT structure, unfolding energetics, and translational control.