A Tale of Two Transitions: The Unfolding Mechanism of the prfA RNA Thermosensor,Biochemistry

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A Tale of Two Transitions: The Unfolding Mechanism of the prfA RNA Thermosensor
Biochemistry ( IF 2.9 ) Pub Date : 2020-11-24 , DOI: 10.1021/acs.biochem.0c00588
Huaqun Zhang ₁ , Ian Hall ₂ , Amos J. Nissley ₂ , Kyrillos Abdallah _{1,

2} , Sarah C. Keane _{1,

2}

Affiliation

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. In Listeria 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 in L. 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.

中文翻译：

两种转变的故事：prfA RNA热传感器的展开机制

在某些细菌信使RNA（mRNA）的5'非翻译区（UTR）中发现的RNA热传感器（RNAT）以依赖温度的方式控制下游基因的翻译。在单核细胞增生性李斯特菌中，关键转录因子PrfA的表达是由RNAT在其5'UTR中介导的。PrfA充当单核细胞增生李斯特氏菌毒力的主要调节剂，控制许多毒力因子的表达。PrfA通过其RNAT元件进行温度调节的表达可作为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，并且先前的研究表明prfA RNAT在狭窄的温度范围内具有高响应能力。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prfA RNAT的展开与cssA（特征明确的RNAT）的展开不同的想法，表明这些RNATs通过不同的机制起作用。我们的数据表明prfA RNAT的展开发生在两个不同的事件中，并且内部环在介导RNAT展开的协同作用中起重要作用。我们进一步证明了核糖体结合位点（RBS）远端的区域不仅有助于RNAT结构的稳定性，而且还影响下游信息的翻译。我们的综合结果提供了有关prfA RNAT结构的热稳定性，展开的能量学和翻译控制的见解。

更新日期：2020-12-08

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